Immunohistochemical Analysis of the Role of Hemocytin in Nodule Formation in the Larvae of the Silkworm,Bombyx mori

Abstract

Hemocytin, a multidomain protein from Bombyx mori L. (Lepidoptera: Bombycidae), is an ortholog of von Willebrand factor and is expected to be a major mediator of hemocyte aggregation. Antiserum was generated against hemocytin, and immune staining of hemocytes, hemolymph, and nodules was performed. Hemocytin was observed in steady-state hemocytes but not in plasma, even after bacterial injection. When hemolymph was smeared on glass slides, hemocytin-containing fibrous structures formed a cellular network mainly consisting of granulocytes and oenocytoids. Hemocytin was stained only in the granules of the granulocytes. When nodule-like aggregates formed 30 sec after bacterial injection, both granulocytes and bacterial cells were observed binding to hemocytin-containing fibrous structures. When nodule sections were stained with antiserum, hemocytin was seen in the matrix of the nodules surrounding the hemocytes. These data suggest that hemocytin plays a major role in nodule formation as a component of the sticky fibrous structure exocytosed from granulocytes.

Mechanisms of nodule-specific melanization in the hemocoel of the silkworm, Bombyx mori

“…5) are released by degranulation and autolysis, respectively, in response to the recognition of microorganisms (Akai and Sato, 1973;Ashida and Brey, 1998). Granulocytes and oenocytoids are major components of nodules during an early stage of their formation (Arai et al, 2013). In this study, 30 min after triggering nodule formation, nodules were composed mainly of central granulocyte aggregates and only a thin outer layer of plasmatocytes (data not shown).…”

“…Fat bodies secrete immune humoral factors, including PRRs, for recognition of invading pathogens, and HPs and SPHs for proPO activation in the hemolymph. Hemocytes are known to play important roles in nodule formation (Arai et al, 2013) and the production of humoral factors, including proPO (Cerenius et al, 2008). To investigate factors involved in nodule melanization in B. mori larvae, we examined the expression patterns of 12 genes encoding PRRs, SPHs, and HPs in fat bodies, hemocytes (the major nodule constituents), and nodules using RT-PCR.…”

“…5e7). Granulocytes and oenocytoids are major constituents of nodules during the early stages of their formation (Arai et al, 2013). Furthermore, granulocyte granules and unidentified molecules in the cytoplasm of oenocytoids may be released by degranulation and autolysis, respectively, in response to microorganism recognition (Akai and Sato, 1973;Ashida and Brey, 1998).…”

“…5e7 and S2), degranulation causes an increase in the concentration of these factors around microorganisms. Other granules contain hemocytin, a putative aggregating factor of hemocytes and microorganisms (Arai et al, 2013). Induced degranulation from granulocytes causes nodule formation, which leads to the aggregation of hemocytes and microorganisms which bound with activated BmSPHs (Figs.…”

Olive fly transcriptomics analysis implicates energy metabolism genes in spinosad resistance

“…vWF tends to bind to other proteins and thus it appears to be involved in multiprotein complexes. In insects, huge vWF-containing proteins, such as hemolectin in D. melanogaster and hemocytin in B. mori , are thought to function in the hemolymph coagulation or hemocyte aggregation processes, such as nodule formation [85, 86]. Such processes are fundamental responses of insect innate immunity in order to clear microorganisms from the hemocoel.…”