A 12R-lipoxygenase in human skin: Mechanistic evidence, molecular cloning, and expression

A recognized feature of psoriasis and other proliferative dermatoses is accumulation in the skin of the unusual arachidonic acid metabolite, 12R-hydroxyeicosatetraenoic acid (12R-HETE). This hydroxy fatty acid is opposite in chirality to the product of the well-known 12S-lipoxygenase and heretofore in mammals is known only as a product of cytochrome P450s. Here we provide mechanistic evidence for a lipoxygenase route to 12R-HETE in human psoriatic tissue and describe a 12R-lipoxygenase that can account for the biosynthesis. Initially we demonstrated retention of the C-12 deuterium of octadeuterated arachidonic acid in its conversion to 12R-HETE in incubations of psoriatic scales, indicating the end product is not formed by isomerization from 12S-H(P)ETE via the 12-keto derivative. Secondly, analysis of product formed from [10R-3H] and [10S-3H]-labeled arachidonic acids revealed that 12R-HETE synthesis is associated with stereospecific removal of the pro-R hydrogen from the 10-carbon of arachidonate. This result is compatible with 12R-lipoxygenase-catalyzed formation of 12R-HETE and not with a P450-catalyzed route to 12R-HETE in psoriatic scales. We cloned a lipoxygenase from human keratinocytes; the cDNA and deduced amino acid sequences share ≤50% identity to other human lipoxygenases. This enzyme, when expressed in Hela cells, oxygenates arachidonic acid to 12-HPETE, >98% 12R in configuration. The 12R-lipoxygenase cDNA is detectable by PCR in psoriatic scales and as a 2.5-kilobase mRNA by Northern analysis of keratinocytes. Identification of this enzyme extends the known distribution of R-lipoxygenases to humans and presents an additional target for potential therapeutic interventions in psoriasis.

Identification and characterization of a retinoid-induced class II tumor suppressor/growth regulatory gene

Retinoids, synthetic and natural analogs of retinoic acid, exhibit potent growth inhibitory and cell differentiation activities that account for their beneficial effects in treating hyperproliferative diseases such as psoriasis, actinic keratosis, and certain neoplasias. Tazarotene is a synthetic retinoid that is used in the clinic for the treatment of psoriasis. To better understand the mechanism of retinoid action in the treatment of hyperproliferative diseases, we used a long-range differential display–PCR to isolate retinoid-responsive genes from primary human keratinocytes. We have identified a cDNA, tazarotene-induced gene 3 (TIG3; Retinoic Acid Receptor Responder 3) showing significant homology to the class II tumor suppressor gene, H-rev 107. Tazarotene treatment increases TIG3 expression in primary human keratinocytes and in vivo in psoriatic lesions. Increased TIG3 expression is correlated with decreased proliferation. TIG3 is expressed in a number of tissues, and expression is reduced in cancer cell lines and some primary tumors. In breast cancer cell lines, retinoid-dependent TIG3 induction is observed in lines that are growth suppressed by retinoids but not in nonresponsive lines. Transient over-expression of TIG3 in T47D or Chinese hamster ovary cells inhibits colony expansion. Finally, studies in 293 cells expressing TIG3 linked to an inducible promoter demonstrated decreased proliferation with increased TIG3 levels. These studies suggest that TIG3 may be a growth regulator that mediates some of the growth suppressive effects of retinoids.

Hyposecretion of the adrenal androgen dehydroepiandrosterone sulfate and its relation to clinical variables in inflammatory arthritis

Hypothalamic–pituitary–adrenal underactivity has been reported in rheumatoid arthritis (RA). This phenomenon has implications with regard to the pathogenesis and treatment of the disease. The present study was designed to evaluate the secretion of the adrenal androgen dehydroepiandrosterone sulfate (DHEAS) and its relation to clinical variables in RA, spondyloarthropathy (Spa), and undifferentiated inflammatory arthritis (UIA). Eighty-seven patients (38 with RA, 29 with Spa, and 20 with UIA) were studied, of whom 54 were women. Only 12 patients (14%) had taken glucocorticoids previously. Age-matched, healthy women (134) and men (149) served as controls. Fasting blood samples were taken for determination of the erythrocyte sedimentation rate (ESR), serum DHEAS and insulin, and plasma glucose. Insulin resistance was estimated by the homeostasis-model assessment (HOMAIR). DHEAS concentrations were significantly decreased in both women and men with inflammatory arthritis (IA) (P < 0.001). In 24 patients (28%), DHEAS levels were below the lower extreme ranges found for controls. Multiple intergroup comparisons revealed similarly decreased concentrations in each disease subset in both women and men. After the ESR, previous glucocorticoid usage, current treatment with nonsteroidal anti-inflammatory drugs, duration of disease and HOMAIR were controlled for, the differences in DHEAS levels between patients and controls were markedly attenuated in women (P = 0.050) and were no longer present in men (P = 0.133). We concluded that low DHEAS concentrations are commonly encountered in IA and, in women, this may not be fully explainable by disease-related parameters. The role of hypoadrenalism in the pathophysiology of IA deserves further elucidation. DHEA replacement may be indicated in many patients with IA, even in those not taking glucocorticoids.