Niacin is an effective lipid-modifying therapy whose use has been limited by suboptimal tolerability. The adverse effect of flushing is due to prostaglandin D2 (PGD2)– mediated cutaneous ...vasodilation. Adjunctive treatment with the PGD2 receptor antagonist laropiprant significantly reduces the incidence and severity of niacin-induced flushing. The objective of this study was to assess the effect of aspirin pretreatment on flushing symptoms with extended-release (ER) niacin/laropiprant in healthy volunteers. A randomized, double-blind, placebo-controlled crossover study compared patient-rated flushing following pretreatment with aspirin 325 mg versus placebo administered 30 minutes before ER niacin 2 g/laropiprant 40 mg. Flushing responses were assessed using participant-reported overall symptom severity score (OSSS), including individual characteristics of redness, warmth, tingling, or itching. The overall incidence and severity of flushing were comparable for participants receiving aspirin or placebo before ER niacin 2 g/laropiprant 40 mg. The difference in 3-day average OSSS between treatments was 0.2 (P = .180). Profiles of flushing severity, frequency, and both-ersomeness were comparable for the aspirin/ER niacin/laropiprant and ER niacin/laropiprant regimens. All treatments were safe and well tolerated. Coadministration of aspirin 325 mg daily with ER niacin 2 g/laropiprant 40 mg does not further reduce residual flushing symptoms associated with ER niacin 2 g/laropiprant 40 mg alone.
Cell cycle withdrawal in postmitotic cells involves cyclin-dependent kinase (Cdk) inhibitors that repress cell cycle Cdk activity. During mouse neurogenesis, cortical postmitotic neurons are shown ...here to accumulate high levels of the p27 Cdk inhibitor compared with their progenitor neuroblasts. Elevated p27 levels in staged embryo brain extracts correlate with p27 binding to Cdk2, and Cdk2 inactivation. Yet, Cdk5, which is associated with the noncyclin activator p35 in neurons, remains active in the presence of high p27 levels. Both in vitro and in vivo, p27 and related inhibitors can recognize a cyclin D--Cdk5 complex but not a p35--Cdk5 complex. The results indicate that the choice of activator determines the susceptibility of Cdk5 to p27 and related Cdk inhibitors, and thus its ability to act in postmitotic cells.
Brain factor-1 (BF-1) is a winged-helix transcription factor which has been shown to be essential for the development of the cerebral hemispheres. We report here the cloning and characterization of ...the mouse BF-1 gene. We have identified two classes of alternatively spliced mouse BF-1 cDNA which are transcribed from distinct promoters. Both classes of cDNA encode identical BF-1 proteins. The class 1 mouse cDNA is the homolog of the previously reported rat and human BF-1 cDNAs, and accounts for about 90–95% of BF-1 expression in brain. Primer extension analyses show that the major promoter, P1, is TATA-less and has multiple transcription start sites. We identified neural cell lines which express BF-1 primarily from the P1 promoter, including the OBL21a and OBL21 cell lines derived from the oldfactory bulb. Expression of BF-1 is highest in proliferating OBL21 cells, declining as the cells differentiate in vitro. This correlates well within the reduction of BF-1 expression as the cells of the telencephalic neuroepithelium differentiate. Using these cells, we demonstrate that the genomic region between −3.7 kb and −79 bp upstream of the P1 promoter contains cell-specific enhancer activity in transient transfection assays.
The genes for rat hepatocyte nuclear factors 3 and 4 (HNF-3 alpha, HNF-3 beta, HNF-3 gamma, and HNF-4) have been mapped in mouse by analysis of restriction fragment length polymorphisms in ...interspecific backcross mice. These hepatocyte-enriched transcription factors are positive-acting transcription factors with binding sites in regulatory regions of many genes expressed in hepatocytes. Both HNF-3 alpha, beta, and gamma and HNF-4 are also expressed in intestine. They have recently been implicated as potential participants in endodermal development from early gut cells because of their close homology to Drosophila genes, which themselves are expressed in the developing gut. Despite having similar functional roles and highly conserved DNA binding domains, the three loci from the Hnf-3 family of genes mapped to three different mouse chromosomes, suggesting that the Hnf-3 family has become widely dispersed during evolution and implying the necessity for independent activation of each member of the HNF-3 family.
The hepatocyte nuclear factor 3 (HNF-3) gene family is composed of three proteins (α, β, and γ) that are transcription factors involved in the coordinate expression of several liver genes. All three ...proteins share strong homology in their DNA binding domains (region I) and are able to recognize the same DNA sequence. They also possess two similar stretches of amino acids at the carboxyl terminus (regions II and III) and a fourth segment of homology at the amino terminus (region IV). Furthermore, the HNF-3 proteins demonstrate homology with the Drosophila homeotic gene fork head in regions I, II, and III, suggesting that HNF-3 may be its mammalian homolog. In order to define HNF-3β protein domains involved in transcriptional activation, we have used a reporter gene, whose transcription is dependent on HNF-3 binding, for hepatoma cell cotransfection assays with expression vectors that produced different truncated HNF-3β proteins. A position-independent activation domain which contained conserved regions II and III was identified at the carboxyl terminus of the HNF-3β protein (amino acids 361 to 458). Moreover, site-directed mutations that altered the sequences within regions H and HI demonstrated their importance to transactivation. The region II-III domain does not possess amino acid sequences in common with other transcription factors and may define a novel activation motif. HNF-3β amino-terminal sequences defined by conserved region IV also contributed to transactivation, but region IV activity required the participation of the region II-III domain. Region IV is abundant in serine amino acids and contains two putative casein kinase I phosphorylation sites, a feature similar to protein motifs described for the transcription factors Pit-1/GHF-1 and HNF-1.
The mouse genomic clone for the prealbumin (transthyretin) gene was cloned, and its upstream regulatory regions were analyzed. The 200 nucleotides 5′ to the cap site when placed within a recombinant ...plasmid were sufficient to direct transient expression in HepG2 (human hepatoma) cells, but this DNA region did not support expression in HeLa cells. The sequence of the 200-nucleotide region is highly conserved between mouse and human DNA and can be considered a cell-specific promoter. Deletions of this promoter region identified a crucial element for cell-specific expression between 151 and 110 nucleotides 5′ to the RNA start site. A region situated at about 1.6 to 2.15 kilobases upstream of the RNA start site was found to stimulate expression 10-fold in HepG2 cells but not in HeLa cells. This far upstream element was invertible and increased expression from the β-globin promoter in HepG2 cells. Unlike the simian virus 40 enhancer, the prealbumin enhancer would not stimulate β-globin synthesis in HeLa cells, and even the simian virus 40 enhancer did not stimulate the prealbumin promoter in HeLa cells. Thus, we identified in the prealbumin gene two DNA elements that respond in a cell-specific manner: a proximal promoter including a crucial sequence between -108 and -151 nucleotides and a distant enhancer element located between 1.6 and 2.15 kilobases upstream.
We previously defined two distinct cell-specific DNA elements controlling the transient expression of the transthyretin gene in Hep G2 (human hepatoma) cells: a proximal promoter region (-202 base ...pairs bp to the cap site), and a far-upstream cell-specific enhancer located between 1.6 and 2.15 kilobases (kb) 5′ of the cap site (R. H. Costa, E. Lai, and J. E. Darnell, Jr., Mol. Cell. Biol. 6:4697-4708, 1986). In this report, we located the effective transthyretin enhancer element within a 100-bp region between 1.96 and 1.86 kb 5′ to the mRNA cap site. In Hep G2 nuclear extracts, three protein-binding sites within this minimal enhancer element were identified by gel mobility and methylation protection experiments. Each binding site was required for full enhancer activity in Hep G2 transient expression assays. Competition experiments in protein-binding assays suggested that two of the three sites were recognized by a similar factor and that the protein interaction with the third site was different. The nuclear protein(s) which bound to the two homologous sites was found mainly or only in cells of hepatic origin, suggesting an involvement of this region in the cell-specific function of this enhancer. The nuclear protein(s) recognizing the third enhancer region was also found in HeLa and spleen cells.
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