Adhesion and cytoplasmic extension of Xylotrechus pyrrhoderus (Coleoptera: Cerambycidae) hemocytes induced by 20-hydroxyecdysone and juvenile hormone in vitro

Insect larval hemocytes change their morphology and function during metamorphosis. However, the mechanisms driving the timing of these changes and the coordination with other premetamorphic events largely remain unclarified. With advancing stages of the beetle Xylotrechus pyrrhoderus prepupal development, the cell density of hemocytes decreased and adherent cells increased when the hemocytes were transferred into basal medium. The response of hemocytes from mature larvae before the prepupal stage to 20-hydroxyecdysone (20E) and juvenile hormone III (JH) was increased adhesion and morphological change; adherent cells accounted for 36.0% and 15.0% of cells in the 20E- and JH-supplemented media, respectively, but only 8.5% of cells in basal medium. Moreover, 60.2% and 53.0% of the adherent cells changed to spindle-shaped cells in 20E and JH treatment, respectively, in contrast with 8.1% in the control. The responses of hemocytes to 20E and JH were dose-dependent, showing stronger responses at concentrations of 10−6 M or more. The extended spindle-shaped cells with longer cytoplasmic extension increased to 30.9% in media supplemented with both 20E and JH, compared with 9.7% for 20E alone and 3.6% for JH alone, suggesting that these two hormones act synergistically. The JH-related compounds, all-trans retinoic acid and farnesol, had no effect. The spindle-shaped cells were also observed at the same level as 20E-treated cells in non-treated cells after the third day of the prepupa stage when ecdysteroid titer is presumably high. These results indicate that the hemocytes of X. pyrrhoderus larvae are responsive to both 20E and JH.