Preprint volume - SIBM

Pre-print Volume - Posters
VARIOUS TOPICS
L. MOLA, V. MINGOLLA, A. GAMBARELLI*, A. PEDERZOLI
Department of Biology, University of Modena and Reggio Emilia,
Via Campi, 213/D – 41100 Modena, Italia.
lucrezia.mola@unimore.it
*Museum of Zoology and Comparative Anatomy, Univ. of Modena and Reggio Emilia, Italia.
IMMUNOREACTIVITY TO HSP70 INDUCTION IN LARVAE
OF DICENTRARCHUS LABRAX (L.) TREATED WITH LPS
INDUZIONE DI IMMUNOREATTIVITÀ ALLA HSP70 IN LARVE
DI DICENTRARCHUS LABRAX (L.) TRATTATE CON LPS
Abstract – The immunoreactivity to HSP70 was tested in 24 day-old larvae of sea bass both controls and
after treatment with LPS. This stress rouses an induction of immunoreactivity in several tissue, such as skin,
gills, gut, liver, renal tubules and hypophysis. The possible role of HSP70 in earlier mechanisms of stress
response was discussed.
Key-words: sea bass, stress, Heat Shock proteins, bacterial antigens.
Introduction – From several years we are carring out immunohistochemical studies
about the appearance and distribution of regolative molecules during the larval stages
of the sea bass, a fish very studied for its commercial value. In previous research (Mola
et al., 2005, 2009; Pederzoli et al., 2007) we hypothesized an active role in the earlier
immunological responses for some well-known molecules involved in responses to
stress, such as ACTH, nitric oxide and CRF. These molecules work before complete
differentiation of lymphocytes associated to gut (GALT), with autocrine/paracrine
ways. In this work we report the results of localization, both in control and in treated
with bacterial antigens animals, of immunoreactivity (IR) to HSP70, a protein of Heat
Shock Protein family, involved in responses to various types of stress.
Materials and methods – Larvae (24 day-old) of Dicentrarchus labrax were treated
with LPS (Lipopolysaccharide from Escherichia coli serotype 055:B5, Sigma). Twenty
of these larvae were maintained in a tank containing 10 U/ml LPS. After 1 h, ten
specimens were immediately fixed, while the remaining ten animals were transferred to
normal tap water for another 1 h and then fixed. All these specimens, together with ten
untreated larvae utilised as controls, were fixed in toto in Bouin fluid, embedded in
paraffin and cut in 7μm transversal sections. Subsequently, the immunocytochemical
procedure was performed. The slides were processed with the biotin-avidin
immunohistochemistry technique (BAS) utilising the polyclonal antibodies from rabbit
anti-Heat Shock protein 70 (HSP70) (d-40) (Santa Cruz, Biotecnology, California,
U.S.A.), titer 1:100.
Results – Untreated specimens showed the presence of a few immunopositive cells in
the skin of the entire body and in the gill epithelium. The apical border of medium and
posterior intestinal epithelium was covered by a thin positive stripe and rare positive
epithelial cells were present in the same regions. A moderate number of positive
hepatocytes and some clusters of positive cells in the hypophysis were observed.
The larvae treated for 1 h with LPS and immediately fixed showed the presence of
many immunopositive cells in the skin and gills. Numerous positive cells were also
41 st S.I.B.M. CONGRESS Rapallo (GE), 7-11 June 2010
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