Search

Lipocalin-type prostaglandin D synthase (L-PGDS) catalyses the formation of prostaglandin D(2) (PGD(2)) and also functions as a transporter for lipophilic ligands, including all-trans retinoic acid (RA). Here, we show that human epidermal melanocytes produce and secrete L-PGDS and PGD(2) in culture medium, whereas L-PGDS is not expressed in human melanoma cell lines, HMV-II, SK-MEL-28, 624 mel and G361. Treatment with RA (1 or 10 microM) for 4 days decreased the proliferation of melanocytes (30% decrease), but not melanoma cells. We therefore isolated L-PGDS-expressing cell lines from 624 mel cells. Treatment with RA decreased the proliferation of L-PGDS-expressing cells by 20%, but not mock-transfected cell lines lacking L-PGDS expression. RA induced expression of a cyclin-dependent kinase inhibitor p21(Cip1) in L-PGDS-expressing cells, but not mock-transfected cells. Moreover, RA increased the transient expression of a reporter gene carrying the RA-responsive elements in L-PGDS-expressing cell lines (at least 5-fold activation), compared to the 2-fold activation in mock-transfected cell lines, suggesting that L-PGDS may increase the sensitivity to RA. Lastly, the knockdown of L-PGDS expression by RNA interference was associated with the restoration of the RA-mediated decrease in proliferation of human and mouse melanocytes. In conclusion, L-PGDS may fine-tune the RA signalling in melanocytes.

Heme oxygenase (HO) catalyzes oxidative breakdown of heme, and constitutes two isozymes, HO-1 and HO-2. Here, we explored the tissue-specific regulation of expression of HO-1 and HO-2 under hypoxemia. There was no significant change in the overall expression levels of HO-1 and HO-2 mRNAs and proteins in the lung during adaptation of C57BL/6 mice to normobaric hypoxia (10% O(2)). However, immunohistochemical analysis revealed the increased expression of HO-1 and HO-2 proteins after 28 days of normobaric hypoxia in the pulmonary venous myocardium that is the extension of the left atrial myocardium into pulmonary venous walls. Moreover, the expression of HO-2 protein was increased in the sub-endocardial myocardium of ventricles under hypoxia, while HO-1 protein level was increased in the full-thickness walls. Thus, hypoxemia induces expression of both HO-1 and HO-2 proteins in the myocardium. Using C57BL/6 mice lacking HO-2 (HO-2(-/-)), which manifest chronic hypoxemia, we also showed that the HO-1 protein level in the lung was similar between HO-2(-/-) mice and wild-type mice. Unexpectedly, HO-1 protein level was lower by 35% in the HO-2(-/-) mouse liver than the wild-type liver. These results indicate that the expression of HO-1 protein is regulated in a tissue-specific manner under hypoxemia.

DESIGN: It has recently been shown that deficiency of adrenomedullin (AM), a potent vasodilator peptide, leads to insulin resistance. We studied expression of AM in NIH 3T3-L1 adipocytes and compared it with expression of resistin, an adipocyte-derived peptide hormone that is proposed to cause insulin resistance. Moreover, we studied the effects of tumor necrosis factor-alpha (TNF-alpha), a known mediator of insulin resistance, on the expression of AM and resistin in 3T3-L1 adipocytes. METHODS: 3T3-L1 cells were induced to differentiate to adipocytes by insulin, dexamethasone and 3-isobutyl-1-methylxanthine. Expression of AM mRNA and resistin mRNA was examined by Northern blot analysis. Immunoreactive AM in the medium was measured by RIA. RESULTS: AM mRNA was expressed in preadipocytes, but barely detectable in adipocytes. Immunoreactive AM was detected in the medium of both preadipocytes and adipocytes, with about 2.5 times higher levels found in preadipocytes. In contrast, resistin mRNA was expressed in adipocytes, whereas it was not detected in preadipocytes. Treatment with TNF-alpha increased AM expression in both adipocytes and preadipocytes, whereas it decreased resistin mRNA levels in adipocytes. CONCLUSIONS: The present study has shown that AM expression was down-regulated and resistin expression was up-regulated during adipocyte differentiation of 3T3-L1 cells. TNF-alpha acted as a potent negative regulator of resistin expression and a potent positive regulator of AM expression in adipocytes, raising the possibility that in addition to its known actions in causing insulin resistance, TNF-alpha may also have actions against insulin resistance through AM and resistin.

The gene coding for microphthalmia-associated transcription factor (Mitf) contains many promoters that could generate multiple Mitf isoforms with distinct amino-termini, such as ubiquitously expressed Mitf-A and Mitf-H. To gain further insight into Mitf isoform multiplicity and the regulation of the promoter usage of the Mitf gene, we have analyzed the function of the amino-terminal domains of Mitf isoforms and the expression of Mitf mRNA in mouse postnatal testis, which is characterized by spermatogenesis and by a cool temperature because of its unique location. Here we show that the amino-terminal domain of Mitf-A possesses a transactivation activity, as judged by yeast expression analysis. We also show the expression of Mitf-A and Mitf-D mRNAs in testis by PCR-based methods. Moreover, in situ hybridization analysis revealed that an Mitf mRNA, probably representing Mitf-A and/or Mitf-D, is expressed in germ cells, including spermatogonia, spermatocytes that undergo meiosis, and round spermatids with the haploid genome, but is undetectable in elongated spermatids with remodeled and condensed chromatin. Notably, Mitf mRNA is undetectable in somatic Leydig cells and peritubular cells. Therefore, multiple promoters may direct differential expression of the Mitf gene in the testis and contribute to functional diversity of Mitf isoforms.