| Atherosclerosis is an inflammatory disease process with increased plasma low density lipoprotein (LDL)-cholesterol level. Monocyte/macrophages contribute to the development of lesions by accumulation of cholesterol esters and the formation of foam cells. The specific atherosclerosis-related genes, e.g. peroxisome proliferator-activated receptor y (PPARy), CD36, and adipocyte fatty-acids-binding protein (aP2), have been investigated for the better understanding of atherosclerotic pathogenesis.PPARy is a nuclear transcription factor and involved in multiple physiological or pathophysio logical processes. In monocyte/macrophages PPARy plays a critical role in regulating expression of CD36 to bind and internalize oxidized LDL (OxLDL). OxLDL, in turn, up-regulates CD36 expression by activatingPPARy. This feed-forward circuit has the potential consequence of perpetuating ever-increasing lipid accumulation and the conversion of macrophages into foam cells. In adipocytes, PPARy crosstalks and cooperates with another transcription factor CCAAT/enhancer binding protein a (C/EBPa) to induce adipocyte differentiation and target genes (e.g. CD36 and aP2) activation. aP2 is highly expressed in adipocytes and serves as shuttles for cytosolic transport and metabolism of fatty acids. aP2 is also expressed by monocyte/macrophages and its deficiency markedly protected hypercholesterolemic (apoE"7") mice from atherosclerosis.In this thesis, we firstly investigate the effect of adipogenic cocktail on macrophage aP2 expression. Adipogenic cocktail consisting of isobutylmethylxanthine (IBMX), insulin (Ins), and dexamethasone (Dex) is routinely used in vitro to induce adipocyte differentiation. Treatment of J774 macrophages with adipogenic cocktail for 4 days considerably induced aP2 mRNA expression. Among the three constituents: Dex was essential and sufficient for the induction of aP2 gene expression, Ins synergized with the effect of Dex, whereas IBMX antagonized the aP2 induction by Dex or Dex plus Ins which was totally opposite to its action in adipocyte differentiation. Consistent with the changes of aP2 mRNA, aP2 protein was increased by Dex that was associated with the enhancement in 14C-oleic acid uptake. The expression of CD36 (increased as aP2 by adipogenic cocktail in adipocytes) was changedinversely to those of aP2 in macrophages. The changes of neither PPARy nor C/EBPa regulated by adipogenic inducers in macrophages were in line with that of aP2 expression. These differences indicate that regulation of macrophage aP2 by adipogenic cocktail and its components may be mediated through signaling pathway independent of PPARy / C/EBPa.Next, we evaluate the effect of tamoxifen (Tarn) on human monocyte/macrophages CD36 expression. Tarn, a routinely used anti-estrogen drug to inhibit breast cancer proliferation, has shown beneficial estrogen-independent cardiovascular protection. The mechanism for this action is so far uncertain. We observed that Tarn and its active metabolite hydroxy-tamoxifen (OH-Tam) decreased CDS 6 protein expression significantly in THP-1 monocytes. The induction of CD36 expression by OxLDL and PGJ2 was also reduced by Tarn or OH-Tam. Parallel reductions in CD36 mRNA levels were observed as measured by RT-PCR or Northern blot. The mechanism study showed that Tarn and OH-Tam down-regulated PPARy expression and simultaneously increased phospho-p44/42 mitogen-activated protein (MAP) kinase mediated phosphorylation of PPARy.Finally, to gain more insight into PPARy we observed the impact of endotoxin lipopolysaccharide (LPS) on PPARy expression in adipocytes, where PPARy is most abundant and its functions are documented well. Treatment of cultured 3T3-Ll adipocytes with LPS resulted in decreased PPARy mRNA and proteinexpression. LPS increased PPARy phosphorylation through activating MAP kinase. Consisting with the changes of PPARy, its target genes, aP2 and CDS 6 expression were also reduced significantly. To adipogenic differentiation LPS inhibited the induction of PPARy protein expression...
|
PDF Full Text Request