CD14 Promoter Research Articles

Promoter region polymorphism of the CD14 gene (C-159T) is not associated with psoriasis vulgaris

Keratinocytes of psoriatic skin show aberrant expression of membrane-bound CD14 (mCD14). In addition, soluble CD14 (sCD14) is elevated in the sera of psoriatic patients. The mechanisms leading to increased CD14 expression and secretion in psoriasis are poorly understood. A bi-allelic polymorphism in the promoter region of the CD14 gene controls CD14 expression on monocytes and sCD14 levels in the sera of healthy subjects. In this context, we explored the CD14 promoter region genotypes of 63 Finnish patients with psoriasis and 126 non-psoriatic controls using a new ARMS-PCR method. No differences in the CD14 genotype frequencies were found between the groups. Thus, our results suggest that the enhanced CD14 expression in psoriasis is not attributable to functional variants of CD14 (-159C/T).

CD14 and TNFα promoter polymorphisms in patients with acute arthritis

Objective. To examine CD14 and TNF f gene polymorphisms in early arthritis in relation to clinical outcome. Methods. We studied 141 Caucasians who had had early arthritis 10 to 38 years earlier. We analysed CD14 (-159) and TNF f (- 238, - 308, - 376) polymorphisms using a novel cycle minisequencing method. DNA pools from 370 Caucasian blood donors served as controls. Results. CD14 (-159)C M T allele frequencies were comparable among patients and controls (39% vs 40%). Fifty men and 42 women had recovered while 24 men and six women had chronic spondyloarthropathy (SpA). Mutant T allele frequency was higher in the chronic SpA group than in the recovered group in women (75% vs 32%, relative risk 1.3, 95% confidence limit 1.1 to 1.6, P= 0.011), but not in men (38% vs 44%). All female patients with chronic SpA had CD14 (- 159)T allele and none had a possibly protective TNF f (- 308)G M A allele. Conclusion. Possession of CD14 (- 159)T allele does not increase risk of ReA but may increase susceptibility of female patients to development of chronic SpA.

A CD14 promoter polymorphism effects soluble CD14 and total IgE levels in infancy

A CD14 promoter polymorphism effects soluble CD14 and total IgE levels in infancy

A common single nucleotide polymorphism in the CD14 promoter decreases the affinity of Sp protein binding and enhances transcriptional activity.

CD14 is a pattern recognition receptor that plays a central role in innate immunity through recognition of bacterial lipoglycans, primarily LPS. Recently, our group has identified a common single nucleotide polymorphism, -159C-->T, in the CD14 proximal promoter. Homozygous carriers of the T allele have a significant increase in soluble CD14, but a decreased total serum IgE. This epidemiologic evidence led us to investigate the molecular basis for the effects of CD14/-159C-->T on CD14 regulation in monocytes and hepatocytes, the two major cell types known to express this gene in vivo. EMSA analysis showed that the T allele results in decreased affinity of DNA/protein interactions at a GC box that contains a binding site for Sp1, Sp2, and Sp3 transcription factors. In reporter assays, the transcriptional activity of the T allele was increased in monocytic Mono Mac 6 cells, which express low levels of Sp3, a member of the Sp family with inhibitory potential relative to activating Sp1 and Sp2. By contrast, both alleles were transcribed equivalently in Sp3-rich hepatocytic HepG2 cells. Our data indicate that the interplay between CD14 promoter affinity and the [Sp3]:[Sp1 + Sp2] ratio plays a critical mechanistic role in regulating transcription of the two CD14 alleles. Variation in a key gene of innate immunity may be important for the pathogenesis of allergy and inflammatory disease through gene-by-gene and/or gene-by-environment interactions.

Promoter polymorphism of the CD14 endotoxin receptor gene as a risk factor for alcoholic liver disease.

Twin concordance studies indicate that genetic factors influence the individual susceptibility for alcoholic liver disease (ALD). Both clinical and experimental data suggest that Kupffer cell activation by gut-derived endotoxins and other bacterial products is an important pathogenic factor. Activated Kupffer cells release proinflammatory cytokines, a process that is regulated by the CD14 endotoxin receptor (CD14). Recently, a C-->T (-159) polymorphism in the promoter region of the CD14 gene was detected and found to confer increased CD14 expression. In the present study, the association of CD14 promoter polymorphism with different forms of ALD was examined in 3 separate autopsy series. Among 442 men with valid alcohol-consumption data, 381 men had been moderate or heavy alcohol consumers. The allele frequency of the CD14 promoter genotype, determined by a modified cycle minisequencing technique, was 0.34 (CC), 0.51 (CT), and 0.16 (TT). The T allele was found to be associated with advanced ALD, i.e., with alcoholic hepatitis (odds ratio [OR]: 2.48; P = .018), and especially with cirrhosis (OR: 3.45; P = .004), but not with fatty liver, periportal fibrosis, or bridging fibrosis. The overall age-adjusted risk for cirrhosis was 3.08 (P = .01) for the carriers of the CT genotype, and 4.17 (P = .005) for the homozygous TT genotype. These results suggest that in the relatively isolated Finnish population, the T allele confers increased risk of alcoholic liver damage. In particular, TT homozygotes are at a high risk to develop cirrhosis.

Polymorphism in the promoter of lipopolysaccharide receptor CD14 and ischemic cerebrovascular disease.

A growing amount of evidence suggests that infectious and inflammatory processes may be involved in the initiation of arteriosclerosis, but the mechanisms are conceivably multifactorial and complex. Two European groups have recently demonstrated that a C(-260)-->T polymorphism in the promoter of the CD14 lipopolysaccharide receptor may be a risk factor for coronary artery disease (CAD). The T allele of this polymorphism reportedly increases the expression of CD14 and may be involved in atherogenesis. In the present study we investigated a possible association between the C(-260)-->T polymorphism in the CD14 promoter and the occurrence of symptomatic ischemic cerebrovascular disease (CVD). Genotype frequencies of the C(-260)-->T polymorphism in the CD14 promoter were determined in 235 patients with CVD, as confirmed by brain CT and/or MRI, and 309 age- and sex-matched control subjects. The distribution of genotypes was as follows: CVD patients, T:/T: 24.3%, C:/T: 53.2%, and C:/C: 22. 6%; controls, T:/T: 26.9%, C:/T: 50.2%, and C:/C: 23.0%. There was no significant difference between the CD14 promoter genotypes of the CVD patients and the controls (chi(2)=0.601, P:=0.741). We also measured the concentration of serum soluble CD14 and the density of membranous CD14 on monocytes in the CVD patients, but the polymorphism was not associated with either the concentration of soluble CD14 or the density of membranous CD14 (P:=0.358, P:=0.238, respectively). Our results indicate that the C(-260)-->T polymorphism in the CD14 promoter is not associated with an increased risk for CVD.

Endotoxins, asthma, and allergic immune responses

Asthma severity depends to a great extent on the levels of endotoxin present in the microenvironment. Although favouring a Th1 cytokine response that could be beneficial to the asthmatic, lipopolysaccharide (LPS) aggravates bronchopulmonary inflammation by several mechanisms. These include neutrophil and eosinophil recruitment, and release by activated macrophages of pro-inflammatory cytokines and nitric oxide. LPS exerts its biological actions through its interaction with CD14. The genetic locus of CD14 is close to the genomic region controlling levels of IgE. A polymorphism in the CD14 promoter region seems to favour high serum IgE levels. Genetic influences may thus control circulating levels of sCD14 and by this mechanism modulate Th1/Th2 balance and IgE synthesis. LPS exposure, although hazardous to the asthmatic, seems to exert a role in the maturation of the immune system in children towards a Th1-skewed pattern.

Common polymorphism in the promoter of the CD14 monocyte receptor gene is associated with acute myocardial infarction in Japanese men

We investigated whether C(-260)-->T polymorphism in the promoter of the CD14 monocyte receptor gene predisposed to coronary atherosclerosis and acute myocardial infarction (AMI) in Japanese men. The frequencies of T allele and T/T homozygotes in patients with AMI were significantly higher than in controls and in patients with angina without prior AMI, suggesting that the CD14 promoter polymorphism is associated with AMI rather than with coronary atherosclerosis, and this polymorphism may be one of the genetic risk factors for AMI in Japanese men.

Characterization of rat CD14 promoter and its regulation by transcription factors AP1 and Sp family proteins in hepatocytes

CD14, a 55 kDa glycoprotein, serves as a lipopolysaccharide (LPS) recognition molecule. CD14 is a monocyte differentiation antigen expressed by myeloid-derived cells, or other cells such as hepatocytes, as either a membrane-bound protein or a soluble serum protein. Increasing evidence indicates that soluble CD14 in plasma is an acute-phase protein derived, among other sources, from liver cells. Although information is available on the cellular expression of CD14, little is known about the cis- and trans-acting factors that regulate basal CD14 transcription in liver cells. We show here that liver cells have a relatively high basal CD14 transcription rate as determined by nuclear run-on assay. We cloned and sequenced an 883 bp 5′-flanking region of the rat CD14 gene and demonstrated functional promoter activity in liver cells. Sequence analysis revealed that, like in the human and mouse CD14 genes, multiple Sp1 and AP1 binding elements exist in rat CD14. Site-directed mutagenesis and transient transfection assays demonstrated that an Sp1 element located at −836 and an AP1 element located at −270 are required for basal promoter activity in liver cells. Electrophoretic mobility shift assays indicate that both Sp1 and Sp3 nuclear factors interact with the −836 Sp1 element, while the AP1-related proteins Fra-2 and JunD bind to the AP1 motif. These data provide novel insights into the regulation of basal CD14 expression in liver cells.

CCAAT/Enhancer-binding Protein Activates the CD14 Promoter and Mediates Transforming Growth Factor β Signaling in Monocyte Development

Transcription factors from the CCAAT/enhancer-binding protein (C/EBP) family play important roles in myeloid cell differentiation. CD14 is a monocyte/macrophage differentiation marker and is strongly up-regulated during monocytic cell differentiation. Here, we report the direct binding of C/EBP to the monocyte-specific promoter of CD14. Transactivation analyses demonstrate that C/EBP family members significantly activate the CD14 promoter. These data indicate that C/EBP is directly involved in the regulation of CD14 gene expression. When myelomonoblastic U937 cells are treated with vitamin D(3) and TGF-beta, they differentiate toward monocytic cells. Using specific antibodies against different C/EBP family members in electrophoretic mobility shift assays and Western blot assays, we have identified a specific increase in the DNA binding and the expression of C/EBPalpha and C/EBPbeta during U937 monocytic cell differentiation, and we found C/EBPalpha and C/EBPbeta bind to the promoter in heterodimer. Furthermore, with stably transfected cell lines, we demonstrate that the C/EBP binding site in the CD14 promoter plays a critical role for mediating TGF-beta signaling in the synergistic activation of CD14 expression by vitamin D(3) and TGF-beta during U937 differentiation. This may indicate that C/EBPs have important functions in the process of TGF-beta signal transduction during monocyte differentiation.

A new promoter polymorphism in the gene of lipopolysaccharide receptor CD14 is associated with expired myocardial infarction in patients with low atherosclerotic risk profile.

Recent findings suggest that inflammation plays a role in atherosclerosis and its acute complications. Cellular response in infections with Gram-negative bacteria is mediated by bacterial lipopolysaccharide (LPS), which activates monocytes to expression of cytokines, growth factors, and procoagulatory factors via LPS receptor CD14. Endothelial cells and smooth muscle cells are stimulated by a complex of LPS and soluble CD14. In this study, LPS receptor CD14 was analyzed to find genetic variants and check them for an association with coronary artery disease or myocardial infarction (MI). When screening the CD14 gene by single-strand conformation polymorphism analysis, a promoter polymorphism was detected and confirmed as a T-to-C exchange at position -159. We determined the genotypes of 2228 men who had undergone coronary angiography for diagnostic purposes. Within the total study group there was no significant association of either genotype with MI or coronary artery disease. However, in a subgroup with low coronary risk (normotensive nonsmokers), a relative risk for MI in probands homozygous for the T allele could be evaluated (OR, 1.6; 95% CI, 1.0 to 2.4; P<0.05). The association was even stronger in low-risk patients older than 62 years (OR, 3.8; 95% CI, 1.6 to 9.0; P<0.01). In conclusion, we describe a new CD14 promoter polymorphism that is associated with MI, especially in older patients with a low atherosclerotic risk profile.

Sp1 is a critical factor for the monocytic specific expression of human CD14

CD14 is a membrane glycoprotein expressed specifically on monocytes and macrophages, and its expression is markedly increased during the process of monocyte differentiation. In order to study CD14 gene regulation, the human CD14 gene was cloned from a partial EcoRI digested chromosome 5 library. A 5.5-kilobase genomic clone contained the full-length CD14 coding sequence and 4.2 kilobases of 5'-upstream sequence. One major and one minor transcription start site were identified 101 and 130 base pairs (bp) upstream, respectively, from the protein translation start ATG. A DNA fragment containing 128 bp of upstream sequence had strong, monocyte-specific promoter activity in the CD14 positive monocytic cell line Mono Mac 6 as compared to the nonmonocytic cell lines HeLa and REX. Four regions in this DNA fragment interact with nuclear proteins isolated from monocytic cells. The Sp1 transcription factor bound to three different regions in the CD14 promoter. Mutation of the major Sp1 binding site (-110 bp) decreased tissue-specific promoter activity, and these results, together with transactivation experiments, demonstrate that Sp1 plays a critical role in the tissue-specific expression of CD14 in monocytic cells. CD14 Sp1 site oligonucleotides bound preferentially to a 105-kDa Sp1 species, which is present in higher relative levels in monocytic than non-monocytic cells, suggesting that modification of Sp1, such as phosphorylation, may explain how the Sp1 site mediates monocytic specific promoter activity.