Guinea-pig primary cell cultures provide a model to study expression and amyloidogenic processing of endogenous amyloid precursor protein

Until now guinea-pigs have been rarely used to investigate formation and de position of Alzheimer's disease-associated amyloid beta peptides despite th e sequence identity of human and guinea-pig amyloid beta peptides being kno wn, and the overall similarity of human and guinea-pig amyloid precursor pr otein. We now describe a primary cell culture system of mixed fetal guinea- pig brain cells, which we have applied to characterize endogenous amyloid p recursor protein processing and amyloid beta formation. These cell cultures were established at embryonic day 24 of guinea-pigs after comparison of se lected stages of guinea pig ontogenetic development with the known ontogeny of rats, and were characterized by immunocytochemical detection of neurona l and glial marker proteins. Amyloid precursor protein expression, processi ng and amyloid beta formation increased in parallel with cellular maturatio n during cultivation and reached a stable phase after approximately 14 days in vitro therefore providing a suitable time for analysis. Aged cultures d isplay strong neuronal amyloid precursor protein immunoreactivity and an al tered profile of amyloid precursor protein isoform messenger RNA expression due to glial proliferation as single neurons were shown to retain their ty pical pattern of amyloid precursor protein expression. We show that amyloid precursor protein in guinea-pig cells is processed by different protease a ctivities which most likely represent alpha- and beta-secretase, leading to the generation of soluble amyloid precursor protein derivatives. Furthermo re, endogenous amyloid precursor protein processing leads to production of substantial amounts of amyloid beta-peptides which accumulate in conditione d culture medium. Amyloid beta was readily detectable by western blot analy sis and was shown to consist of approximately 80-90% amyloid beta(1-40). We suggest that primary guinea-pig cell cultures provide a valuable tool in amyloid research that resembles amyloid precursor protein processing under physiological concentrations and, therefore, the situation in humans more closely than current rodent models. It should be especially useful in scree ning experiments for secretase inhibiting compounds. (C) 1999 IBRO. Publish ed by Elsevier Science Ltd.