Giridhar, J. and D. Acosta. Elevation of Intracellular Calcium by 13-Cis Retinoic Acid (13-Cis RA) in Primary Rat Keratinocyte Model: Implications for Toxicity. The Toxicologist. February 1992. 12(1): 297. (Abstract #1146).

cis-retinoic acid - 4759-48-2; trans-retinoic acid - 302-79-4

Topical application of retinoids may cause mucocutaneous side effects. The mechanism of retinoid induced dermal side effects, such as severe skin irritation and scaling, has not been thoroughly investigated. Primary rat keratinocytes, cultured from neonatal rat pups, provide a good in vitro model to study the mechanism of retinoid toxicity. 3-4 day old keratinocyte cultures grown on glass coverslips were loaded with 1 uM fura-2AM, in MCDB 153 medium, for 30 min at 37 C, washed with Ca-Krebs Ringer buffer and evaluated by digitized fluorescence imaging. Cells were incubated with different concentrations of 13 cis-RA (10(-10) M to 10(-4) M) for 4 and 24 hr, and at the end of the treatment period, cells were loaded with fura-2AM and intracellular calcium was measured. The 4 and 24 hr treatments with 13-cis RA did not appear to alter intracellular calcium levels. In contrast, cells which were loaded initially with fura-2AM and then treated for 30 min with RA [10(-10) M] showed a sustained increase in intracellular calcium, indicated by a 2 fold sustained increase in intracellular calcium. Integrity of cell membranes was monitored by propidium iodide fluorescence technique. 10(-4) M RA at the end of the 24 hr was toxic to cells resulting in loss of cell viability. Integrity of the cell membrane was also evaluated by measuring the leakage of LDH into the media. Significant increases in LDH release after 30 min, 4 and 24 hr periods were seen only at 10(-4) M RA. These results indicate a role for calcium in RA induced dermatotoxicity.