DEVELOPMENT OF AN IN VITRO RAT NASAL EPITHELIAL MODEL FOR PREDICTING UPPER RESPIRATORY TRACT TOXICITY

Abstract

Nasal toxicology is an important, expanding area of research, which has, in the past, centered on in vivo studies. There is now a need for in vitro methodology to study this target organ, and the aim of this work was to develop such a model using isolated rat nasal turbinates. Initially, optimal culture conditions for olfactory epithelium (OE)and respiratory epithelium (RE) were determined using lactate dehydrogenase (LDH) release as an indicator of viability. The optimal culture conditions were found to be incubation at 31 C under an atmosphere of 95%O:5%CO2, in six-well plates containing 2 mL of William's E medium supplemented with glutamine (2 mM) and gentamycin (100 mug/mL), with shaking at 60 strokes/min (amplitude 3.5 cm). Under these conditions, LDH release from both tissues was 30-40% by 24 h. The model was further characterized in terms of a rangeof viability parameters reflecting different aspects of cellular function. Initial values for intracellular potassium, ATP, and nonprotein sulfhydryl levels were 1 mumol/mg protein, 4.43 nmol/mg protein, and 33 nmol/mg protein, respectively, for OE, and 0.84 mumol/mg protein, 5.53 nmol/mg protein, and 31 nmol/mg protein, respectively, for RE. During a 24-h incubation period these parameters did not fall significantly below these initial values in either tissue. Protein synthesis was maintained throughout the24-h incubation, and the rate of synthesis in RE was double that of OE. Histological examination of the tissues demonstrated that RE maintained normal morphology for 24 h, but OE showed signs of focal degeneration as early as 4 h after the start of the incubation period. Thus, RE was maintained in a viable state for 24 h, but OE for only 8 h.

Keywords: :

• In Vitro Nasal Cavity Olfactory Epithelium Respiratory Epithelium