Steven Ewart with Andrea Romani (SOM, Dept. of Physiology & Biophysics)

Role of Cellular Mg2+ in Modulating Collagen Deposition and Interleukin Release in Ito and Kupffer Cells

Introduction: Magnesium ion (Mg2+) is the second most abundant cation after potassium within mammalian cells, including liver cells. At the cellular level, Mg2+ has been reported to regulate more than 180 enzymes, especially those involved in cell cycle regulation and glucose metabolism. Acute and chronic ethanol administration reduces total Mg2+ content in liver tissue. A decrease in cellular Mg2+ content has been associated with increased pro-inflammatory cytokines release and altered collagen deposition.

Hypothesis: In the present study I will test the hypothesis that a reduced cellular Mg2+ content within Kupffer and Ito (stellate) cells is sufficient to generate an increased production of Il-2 and Il-6 and an increased collagen deposition, respectively, or it predisposes to these changes in the presence of ethanol administration.

Methods: Kupffer and Ito cells isolated from rat livers will be maintained in culture in the presence of varying extracellular Mg2+ concentrations (i.e. 0.4, 0.8 and 2 mM). Cell cultures will be challenged by addition of different doses of ethanol ranging from 0.01% to 1% for varying period of time spanning from 15 min to 96 hours. Cellular Mg2+ content will be measured by atomic absorbance spectrophotometry (AAS). Release of IL-2 and IL-6 in the extracellular medium will be assessed by Elisa assay. Collagen deposition will be determined by 3H-proline incorporation. mRNA levels for Il-2, Il-6 and collagen will be determined by RT-PCR.