The specific molecular mechanisms and nuclear events involved in leading to the transcription of pathological growth factor and inflammatory genes under diabetic conditions are not fully resolved. Circulating monocytes in diabetic individuals would be continuously exposed to hyperglycemic conditions. Monocyte activation, adhesion, transmigration and foam cell formation are key events in the pathogenesis of atherosclerosis. There is evidence of increased leukocyte-endothelial interactions in diabetic animals and with monocytes from human diabetic subjects. However, the behavior of monocytes cultured under high glucose (HG) and diabetic conditions have not been well studied. Much less is known about in vivo transcription mechanisms leading to the regulation under diabetic conditions of inflammatory cytokine and chemokine genes such as tumor necrosis factor- alpha (TNF-alpha), monocyte chemoattractant protein-1 (MCP-1) and cyclooxygenase-2 (COX-2) that are implicated in monocyte activation and atherosclerosis. The hypothesis is that diabetic conditions lead to in vivo nuclear chromatin remodeling and key alterations in the nuclear transcriptome machinery. This induces the transcription of NF-kB-regulated inflammatory genes in monocyte/macrophages and leads to enhanced monocyte activation and adhesion.
The Specific Aims are: 1. To determine whether HG conditions increase interactions between NF-kB p65 transcriptionally active subunit and key transcriptional coactivators with histone acetyl transferase activity in monocytes. 2. To examine in vivo nuclear transcription and novel chromatin remodeling mechanisms by which HG and advanced glycation end products (AGEs) lead to inflammatory gene transcription in monocytes. 3. To evaluate the functional relevance of NF-kB activation under HG and AGE treated conditions by adopting state-of-the-art RNA-interference techniques to silence NF-kB p65. The project is supported by our preliminary data that human monocytes under HG conditions, as well as monocytes from diabetic patients produce significant amounts of TNF-alpha and MCP-1 in an oxidant stress and NF-kB-dependent manner. Our new data also shows evidence of chromatin remodeling under HG conditions. This project adopts several innovative approaches including in vivo chromatin screening and gene silencing, and could lead to novel new therapies for diabetic complications. ? ?
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