Evidence that P2 purinergic signaling in astrocytes regulates expression of N-cadherin and thrombospondin-1: Implications for CNS injury and development
Date of Award
Doctor of Philosophy (Ph.D.)
First Committee Member
Ellen Barrett - Committee Chair
Purinergic regulation of N-cadherin and thrombospondin-1 (TSP-1), molecules produced by astrocytes that are involved in cell-cell and cell-matrix interactions, is unknown. I used primary cultures of rat cortical astrocytes to study the effects of purinergic signaling on the expression of N-cadherin and TSP-1. N-cadherin and TSP-1 protein expression increased in a time- and concentration-dependent manner with extracellular ATP. Cultures treated with a P1 antagonist, 8-(p-sulfophenyl)-theophylline, or P2 antagonists, reactive blue 2 or pyridoxalphosphate-6-azophenyl-2'-4'-disulfonic acid (PPADS), showed reduced ATP-induced expression of N-cadherin and TSP-1, suggesting P1 and P2 receptors involvement in the regulation of N-cadherin and TSP-1 expression. However, agonists studies showed that UTP and 3'-O-(4-benzoyl)benzoyl-ATP (BzATP), but not 2-methylthioADP (2MeSADP), adenosine, and N6-cyclopentyladenosine, statistically increased N-cadherin expression, while UTP but not BzATP, 2MeSADP, adenosine or N-ethylcarboxamidoadenosine statistically increased TSP-1 expression, suggesting that ATP-induced N-cadherin expression is mediated by both P2Y and P2X receptors while ATP-induced TSP-1 expression is mediated by P2Y receptors. Using various protein kinase pathway inhibitors, I found that the ATP-induced N-cadherin expression involves both extracellular signal-regulated protein kinase (ERK) and protein kinase B/Akt. However, other signaling pathways may also be involved since inhibition of both ERK and Akt did not completely block the ATP-induced N-cadherin expression. I found that ERK, Akt, p38-mitogen-activated protein kinase (MAPK) and c-Jun N-terminal protein kinase/stress activated-protein kinase were all involved in the ATP-induced TSP-1 expression, particularly p38/MAPK and Akt. Using an in vitro model of CNS mechanical injury, N-cadherin and TSP-1 expressions increased in a time-dependent manner after mechanical strain. Injury-induced TSP-1 expression involves P2 receptor activation since PPADS was able to block the injury-induced TSP-1 expression. In vivo studies with moderate traumatic brain injury showed that the expression of various P2 receptors changes with injury, particularly P2X 1 and P2X7 with increased mRNA but decreased protein levels. I have identified the signaling mechanisms involved in regulating N-cadherin and TSP-1 expression in rat astrocytes. These studies suggest that purinergic signaling may be an important factor in N-cadherin and TSP-1 mediated cell-matrix and cell-cell interactions such as occur during development and after CNS injury.
Tran, Minh D., "Evidence that P2 purinergic signaling in astrocytes regulates expression of N-cadherin and thrombospondin-1: Implications for CNS injury and development" (2005). Dissertations from ProQuest. 2217.