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Gut chitinase, originally identified as acidic mammalian chitinase, is secreted from the salivary gland and stomach in humans and mice, but exclusively from the liver in cattle. Since the organs producing gut chitinase differ depending on the species, here we determined the expression site of this enzyme in the gastrointestinal tract of frogs, Xenopus Iaevia.-: and Rana caresbeiuna, which eat a large amount of chitin-coated animals, using in sift: hybridization and immunohistochemistly. The chitinase was detected only in the oxynticopeptic cells of gastric glands and was accumulated in secretoly granules in the cytoplasm, suggesting that the enzyme may be released into the gastric lumen directly. In addition, the enzyme was not found in the stomach of larvae, but was detected at stage 62 of metamolphosis and later, when the frogs changed from a herbivorous to a carnivorous diet. Chitin is the most abundant glycopolymer i11 the integument of many species, such as insect exoskeletons, shells of crustaceans, and fungal cell walls (4, ll). Chitin-fragmenting hydrolases, chitinases, are also expressed by a vast array of organisms, including even those that do not contain chitin in the body (3). It was long generally believed that man and other mammals did not possess an enzyme analogous to chitinase (5, 9). However, Boot er cl. (2) demonstrated that human macrophages produce and secrete chitinase (chitotriosidase). They also found the presence of a second mammalian chitinase, referred to as acidic mammalian chitinase (AMCase), Corresponding to: Dr. Toshihiko Iwanaga, Laboratory of Anatomy, Graduate School of Veterinary Medicine, Hokkaido University, Kita I8-Nishi 9, Kita-ku, Sapporo 0600813, Japan Tel: +8l-ll-F06-5187 Fax: +81-ll-"F06-5189 E-mail: tiwanaga@vetmed.hokudai.ac.jp *Present address: Cellular Biochemistry, Animal Resource Science, Veterinary Medical Science, The University of Tokyo, l—l-l Yayoi, Bunkyo-ku, Tokyo 1138657, Japan in the salivary gland and the stomach of man and rodents (I). We recently found the presence of a chitinbinding protein (CBPbO4) in bovine serum and identified it as an AMCase homologue. Interestingly, the bovine chitinase is only expressed in the hepatic parenchymal cells (12). In addition, we identified chicken AMCase (CBPch04) which is expressed in both the stomach and liver, and thus proposed calling this type of enzyme “gut chitinase” (13). These mammalian and avian gut chitinases are highly homologous to each other, with more than 75% amino acid sequence homology. Gut chitinase may be one of the most important enzymes in animals that eat chitin-coated animals, such as insects. Recently, Oshima er al. (8) have cloned a putative gastric chitinase cDNA from the toad stomach as a counterpart of mammalian gut chitinase. However, the precise production site of the gut chitinase in the anura and its developmental expression pattern remain unknown. In this study, we examined the cellular and subcellular localization of gut chitinase in two frog species, Xenopus laevis and Rana caresbeiano by in sire: hybridization and imntunohistochemical analysis, and also exam-

Cellular Expression of the Gut Chitinase in the Stomach of Frogs Xenopuslaevis and Rana catesbeiana