Novel cysteine-rich secretory protein in the buccal gland secretion of the parasitic lamprey, Lethenteron japonicum.

Lampreys are one of the most primitive vertebrates diverged some 500 million years ago. It has long been known that parasitic lampreys secrete anticoagulant from their buccal glands and prevent blood coagulation of host fishes. We found two major protein components of 160 and 26 kDa in the buccal gland secretion of parasitic river lamprey, Lethenteron japonicum. The larger protein was identified as river lamprey plasma albumin. The complete primary structure of the 26-kDa protein was determined by protein and cDNA analysis. It belonged to the cysteine-rich secretory protein (CRISP) superfamily that includes recently identified reptile venom ion-channel blockers. Lamprey CRISP blocked depolarization-induced contraction of rat-tail arterial smooth muscle, but showed no effect on caffeine-induced contraction. The result suggests that lamprey CRISP is an L-type Ca(2+)-channel blocker and may act as a vasodilator, which facilitates the parasite to feed on the host's blood. The lamprey CRISP protein contains a number of short insertions throughout the sequence, when aligned with reptilian venom CRISP proteins, probably due to the large evolutionary distance between the Agnatha and the Reptilia, and may represent a novel class of venom CRISP family proteins.

[Possible involvement of Ankhzn, a novel protein possessing FYVE domain, in cellular endocytosis and autophagocytosis in vitro].

The objective of this study was to investigate the functional significance of Ankhzn, a novel zinc binding FYVE finger protein, which was previously identified by our gene trap experiment. On the basis of the fact that a family of FYVE double zinc finger proteins is involved in membrane transport, the present immunohistochemical study was undertaken with a focus on the localization of Ankhzn and endocytotic structures. First, the distribution of Ankhzn was observed in mouse macrophages that were in the process of incorporation of dextran particles in vitro. Ankhzn was further demonstrated in phagocytosed transferrin labeled with Texas red. Second, the topological relationship between the localization of Ankhzn and that of transferrin receptor was studied in neuroblastoma-derived B103 cells. Some but not all Ankhzn immunoreactive vesicles simultaneously contained transferrin receptor. Third, the immunoreactivity of Ankhzn was compared between serum-supplemented and serum-free cultures of B103 cells. A marked increase in the immunostaining of Ankhzn was seen in starved B103 cells. All of these observations suggest that Ankhzn is essential for the formation of endocytotic and autophagocytotic vesicles.

Immunohistochemical study on the fetal rat pituitary in hyperthermia-induced exencephaly.

Hyperthermia of fetal rats is known to cause malformations of various organs including brain. The present study was carried out to investigate the effect of the hyperthermia-induced brain damages on the development of the adenohypophysis. Mother rats of day 9.5 of pregnancy were anesthetized and immersed in hot water (43 degrees C) for 15 min. At day 21.5 of gestation, fetuses were removed by caesarian section and examined for exencephaly. Hyperthermal stress induced varying degrees of exencephaly in 36% of surviving fetal rats. In extreme cases a considerable part of head was lost. Even in those fetuses with severe brain deformities, the hypophysial stalk and neural lobe were present though they were markedly underdeveloped. In exencephalic fetuses, no immunoreactive vasopressin was detected in the neural lobe of the hypophysis. Immunohistochemical examination of the adenohypophysis showed that exencephaly caused a marked decrease in the number of growth hormone (GH)-producing cells. Other types of hormone-producing cells appeared to be unaffected by brain anomaly. The reason for a decreased population of GH cells in exencephalic fetuses is discussed in relation to their adrenocortical hypotrophy.

Differentiation of Striated Muscle Fibers in Monolayer Cultures of Rat Anterior Pituitary: (rat pituitary/cell culture/differentiation/striated muscle).

Striated muscle fibers appeared in monolayer cultures of rat anterior pituitary cells maintained in αMinimum Essential Medium (αMEM). As muscle differentiation in cultures of pituitary cells under ordinary conditions has not hitherto been reported, an in vitro study was undertaken to determine what factor(s) is responsible for this myogenesis. When dispersed anterior pituitary cells were culrured in three different media, αMEM, Medium 199 and Dulbecco's Modified Eagle Medium (DMEM), only αMEM induced a high incidence of striated muscles. The nature of the serum (fetal calf, calf and horse) and its concentration (1-10%) did not affect myogenesis. In monolayers in αMEM, the sequence of differentiation of striated muscle was as follows: 1) Elongated cells, resembling myoblasts appeared; 2) these cells fused; and finally 3) cross striations appeared. Rhythmic contraction was most intense in striated muscle fibers, but it was also obsrved in myotubes without cross striations and even in myogenic cells before fusion. The possible origin of muscles in these pituitary cultures is discussed.