Vascular Cell Adhesion Molecule-1 (VCAM-1): Molecular Biologic and - Pathologic Studies of Its Role As Vascular and Mesangial Inflammatory Marker
Posted on: Wednesday, 15 September 2004, 06:00 CDT
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Aims: This study intended to determine the structure and function of VCAM-1 as vascular and mesangial inflammatory marker gene.
Methods: various molecular (cloning, sequencing, electromobility shift assay (EMSA), transfection, primer extension, qualitative and quantitative RT-PCR), cell culture and histopathological (laser- microdissection, immunohistology) techniques and animal models
Results: The structure of human VCAM-1 was determined from a human endothelial library and its rat homologe cloned by PCR using degenerative primers and RNA from lung tissue of LPS treated rats. The temporal and spatial expression was defined in a mouse model of endotoxemia. Using quantitative RT-PCR, the temporal expression was quantitated in human mesangial, venous and microvascular endothelium after TNF alpha stimulation. The transcriptional regulation of VCAM- 1 via the p65/p50 complex of NF-[kappa]B was demonstrated by EMSA in renal mesangial cells. NF-[kappa]B acticvation could be up-/ downregulated in vitro and in vivo by modifying the levels of gluthathione, leading to high or non-detectable VCAM-1 expression levels. The potential use of VCAM-1 as marker of inflammation was determined in paraffinized human renal biopsies of different glomerular lesions after laser-microdissection by quantitative RT- PCR. In particular, VCAM-1 levels were upregulated in mesangial- proliferative/IgA-nephropathy, thought to be a non-inflammatory disease.
Conclusions: VCAM-1 can serve as positive marker gene of inflammatory, NF-[kappa]B mediated gene expression for future microarray-based experiments to identify the complexity of signaling and gene expression in inflammation.
J.W.U. FRIES
Institut fr Pathologie, Universitt Kln
Copyright Urban & Fischer Verlag 2004
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