Aspirin inhibits tumor necrosis factor-alpha-stimulated fractalkine expression in human umbilical vein endothelial cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Fractalkine is an important chemokine mediating local monocyte accumulation and inflammatory reactions in the vascular wall. Aspirin inhibits inflammatory cytokine expression closely related to atherosclerosis through the way independent of platelet and cyclooxygenase (COX). There has been no report about the effect of aspirin on fractalkine expression. We aimed to determine the fractalkine expression in human umbilical vein endothelial cell (HUVEC) stimulated by tumor necrosis factor (TNF)-alpha and the effect of aspirin intervention.</p><p><b>METHODS</b>Six of 8 HUVEC groups received either different concentrations of aspirin (0.02, 0.2, 1.0, 5.0 mmol/L) or 40 micromol/L pyrrolidinecarbodithioc acid (PDTC) or 0.5 micromol/L NS-398. The other two groups were negative control and positive control (TNF-alpha-stimulated). After being incubated for 24 hours, cells of the 8 groups except the negative control one were stimulated with TNF-alpha (4 ng/ml) for another 24 hours. After that, the cells were collected for RNA isolation and protein extraction.</p><p><b>RESULTS</b>Both mRNA and protein expressions of fractalkine in HUVEC were upregulated by 4 ng/ml TNF-alpha stimulation. Aspirin inhibited fractalkine expression in a dose-dependent manner at mRNA and protein levels. Nuclear factor-kappa B inhibitor, PDTC, effectively decreased the fractalkine expression. Fractalkine expression was not influenced by COX-2 selective inhibitor NS-398. COX-1 protein expression was not changed by either TNF-alpha stimulation or aspirin, PDTC, NS-398 intervention. Both mRNA and protein expression of COX-2 in HUVEC were upregulated by 4 ng/ml TNF-alpha stimulation. Aspirin decreased COX-2 expression in a dose-dependent manner at mRNA and protein levels.</p><p><b>CONCLUSIONS</b>TNF-alpha-stimulated fractalkine expression is suppressed by aspirin in a dose-dependent manner through the nuclear factor-kappa B p65 pathway.</p>

Apoptosis induced by short hairpin RNA-mediated STAT6 gene silencing in human colon cancer cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>The relationship between signal transduction and tumors has become one of the foci in cancer research. Signal transducer and activator of the transcription 6 (STAT6) signaling pathway is found to be activated in some cancer cells. But the function of the pathway in cancer cells is unknown. This study was undertaken to investigate the effect of the Stat6 signaling pathway on apoptosis in human colon cancer cells (HT-29 cells) and the possible mechanism of Stat6 by RNA interference techniques.</p><p><b>METHODS</b>Four eukaryotic expression plasmid vectors of short hairpin RNA (shRNA) specific for the STAT6 gene were designed and generated by molecular biological technology. The plasmid vectors were transfected into HT-29 cells by cation liposomes to block the Stat6 signaling pathway. The expressions of STAT6 mRNA and phosph-Stat6 protein were detected by the reverse transcriptase polymerase chain reaction (RT-PCR) method and flow cytometry respectively to screen the most effective shRNA at 72 hours after transfection. The apoptosis condition of the cells in which the expression of the STAT6 gene had been interfered was analyzed by flow cytometry and confocal microscopy. Both mRNA and protein expression of B cell lymphoma-2 (Bcl-2) and Bax were detected by RT-PCR and western blotting.</p><p><b>RESULTS</b>Two effective eukaryotic expression plasmid vectors of shRNA specific for the STAT6 gene were generated successfully. One can reduce the expression of the STAT6 gene by 82.4% and the other by 56.8% (P < 0.01). The apoptotic rate of colon cancer cells in which STAT6 gene expression had been interfered was significantly higher than that in controlled colon cancer cells (P < 0.01). In the cells in which the Stat6 signaling pathway was blocked, the levels of mRNA and protein Bcl-2 were significantly decreased, whereas those of Bax were significantly increased (P < 0.01).</p><p><b>CONCLUSIONS</b>The Stat6 signaling pathway can inhibit apoptosis in human colon cancer cells. The subsequent disorder of Bcl-2/Bax expression may play an important part in that process. The STAT6 gene may serve as a potential target in cancer therapy.</p>

Mechanism linking atherosclerosis and type 2 diabetes: increased expression of scavenger receptor CD36 in monocytes / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>We investigated the pathogenesis of atherosclerosis in diabetes, and detected the expression of scavenger receptor CD36 in monocytes in patients with type 2 diabetes.</p><p><b>METHODS</b>According to the criteria by WHO, diabetic patients were classified into two groups: well controlled diabetic patients (WCP) and poorly controlled diabetic patients (PCP). The expression of CD36 protein and mRNA were evaluated by flow cytometry and reversal transcription polymerase chain reaction (RT-PCR). Plasma levels of accumulation of oxidized LDL (oxLDL) were directly measured by sandwich enzyme-linked immunosorbent assay (ELISA) method.</p><p><b>RESULTS</b>Flow cytometry and RT-PCR showed that the mean fluorescence intensity (MFI) of CD36 in monocyte and CD36 mRNA were significantly higher in the PCP and WCP in comparison with healthy controls (P<0.01). CD36 MFI and mRNA in the PCP were increased by 78% and 36% compared to the WCP. In both groups, CD36 MFI and mRNA were significantly higher in patients with diabetic atherosclerosis in comparison with those without diabetic atherosclerosis (P<0.05). No significant difference was found in CD14 expression between the groups (P>0.05). The concentrations of plasma oxLDL were higher in the PCP group compared to WCP and control group (P<0.05), whereas oxLDL average values did not differ significantly between WCP and control groups (P>0.05). In the WCP and PCP groups, oxLDL levels were higher in patients with diabetic atherosclerosis than those without diabetic atherosclerosis (P<0.05).</p><p><b>CONCLUSIONS</b>The increased expression of scavenger receptor CD36 may be one of the mechanism of accelerated atherosclerosis in diabetic. The poorly controlled diabetes patients are at higher risk for the vascular complications than the well controlled diabetic patients.</p>