Thromboxane Mobilizes Insect Blood Cells to Infection Foci

Abstract

Innate immune responses are effective for insect survival to defend against entomopathogens including a fungal pathogen, Metarhizium rileyi, that infects a lepidopteran Spodoptera exigua. In particular, the fungal virulence was attenuated by cellular immune responses, in which the conidia were phagocytosed by hemocytes (insect blood cells) and hyphal growth was inhibited by hemocyte encapsulation. However, the chemokine signal to drive hemocytes to the infection foci was little understood. The hemocyte behaviors appeared to be guided by a Ca²⁺ signal stimulating cell aggregation to the infection foci. The induction of the Ca²⁺ signal was significantly inhibited by the cyclooxygenase (COX) inhibitor. Under the inhibitory condition, the addition of thromboxane A₂ or B₂ (TXA₂ or TXB₂) among COX products was the most effective to recover the Ca²⁺ signal and hemocyte aggregation. TXB₂ alone induced a microaggregation behavior of hemocytes under in vitro conditions. Indeed, TXB₂ titer was significantly increased in the plasma of the infected larvae. The elevated TXB₂ level was further supported by the induction of phospholipase A₂ (PLA₂) activity in the hemocytes and subsequent up-regulation of COX-like peroxinectins (SePOX-F and SePOX-H) in response to the fungal infection. Finally, the expression of a thromboxane synthase (Se-TXAS) gene was highly expressed in the hemocytes. RNA interference (RNAi) of Se-TXAS expression inhibited the Ca²⁺ signal and hemocyte aggregation around fungal hyphae, which were rescued by the addition of TXB₂. Without any ortholog to mammalian thromboxane receptors, a prostaglandin receptor was essential to mediate TXB₂ signal to elevate the Ca²⁺ signal and mediate hemocyte aggregation behavior. Specific inhibitor assays suggest that the downstream signal after binding TXB₂ to the receptor follows the Ca²⁺-induced Ca²⁺ release pathway from the endoplasmic reticulum of the hemocytes. These results suggest that hemocyte aggregation induced by the fungal infection is triggered by TXB₂via a Ca²⁺ signal through a PG receptor.

Keywords: Spodoptera exigua; fungi; hemocyte; insect; thromboxane.