Physiological evidence of integrin‐antibody reactive proteins influencing the innate cellular immune responses of larval Galleria mellonella hemocytes

Larval Galleria mellonella (L.) hemocytes form microaggregates in response to stimulation by Gram‐positive bacteria. Hemocyte adhesion to foreign materials is mediated by the cAMP/ protein kinase A pathway and the β‐subunit of cholera toxin using a cAMP‐independent mechanism. Cholera toxin‐induced microaggregation was inhibited by the integrin inhibitory RGDS peptide, implying integrins may be part of the mechanism. Based on the types of mammalian integrin‐antibody reactive proteins affecting hemocyte adhesion and bacterial‐induced responses α₅, α_v, β₁, and β₃ subunits occurred on both granular cell and plasmatocyte hemocyte subtypes. A fluorescent band representing the binding of rabbit α₅‐integrin subunit antibodies occurred between adhering heterotypic hemocytes. The frequency of the bands was increased by cholera toxin. The α₅ and β₁ rabbit integrin subunit antibodies inhibited removal of Bacillus subtilis (Cohn) from the hemolymph in vivo. A α₅ β₁‐specific synthetic peptide blocker similarly diminished hemocyte function whereas the α_v β₃‐specific inhibitory peptide and the corresponding integrin subunit antibodies did not influence nonself hemocyte activities. Western blots revealed several proteins reacting with a given integrin‐antibody subtype. Thus integrin‐antibody reactive proteins (which may include integrins) with possible α₅ and β₁ epitopes modulate immediate hemocyte function. Confocal microscopy established plasmatocyte adhesion to and rosetting over substrata followed by granular cell microaggregate adhesion to plasmatocytes during early stage nodulation.