Previous posts discussing infection management in
aquaculture have referred to the paradigm that invertebrates do not have acquired
immune systems like us vertebrates. Without antibodies, how does the
invertebrate immune system rely completely on innate (non-specific) defences? This
study details an interesting mechanism of how the innate immune system responds
to pathogens.
Litopenaeus vannamei is a shrimp widely devoured by Homo sapiens
and is assumed to have an innate immune system, defending the arthropod using melanisation,
hemolymph clotting, foreign particle trapping and good old phagocytosis. The
hemolymph enzyme transglutaminase (TGase) is instrumental in all of these
processes and inhibiting its production in shrimp results in bacteria rich
hemolymph and very sick crustaceans. Hemocytes produce prophenoloxidase
(proPOase), the precursor to another important immune enzyme, phenoloxidase (POase).
Once activated, PO facilitates phenol oxidation to antimicrobial quinones, which
can also polymerize into melanin to encapsulate pathogens. Basically, TGase and
proPO are what makes the crustacean innate immune system tick.
As you know, heat shock proteins (HSPs) are a highly
multifunctional family of proteins used by all organisms. The family HSP70 is
the most highly conserved and studied; recently it has been flagged as a
notable regulator in the initial innate immune system response, improving POase
activity in Artemia infected with Vibrio. But such studies are controversial;
many have used a recombinant HSP70 extracted from E. coli called DnaK, which may have been contaminated with endotoxins
able to activate the immune system in a similar way.
This study aimed to determine whether or not HSP70 homologue
DnaK induces immune response in L. vannamei,
by measuring changes in the expression of proPOase and TGase genes in response
to an injection of either recombinant HSP70 (DnaK) from E. coli or a
contaminant-free control of chemically synthesised HSP70 (synDnaK). All shrimp
used were specific pathogen free (SPF) and given a high and a low dose
injection. Gene expression was measured by synthesising cDNA from isolated RNA,
then amplifying it by quantitative PCR with TGase and proPOase primers. TGase
gene expression increased very significantly in response to low and high doses
of DnaK, low doses of synDnaK, but did not change for high dose synDnaK.
Expression of a proPOase gene increased significantly for low and high doses of
DnaK; a similar pattern was observed for synDnaK. DnaK contamination was
measured also, showing endotoxin levels much lower than concentrations found to
induce crustacean immune response in other studies.
Recombinant DnaK and synthetic DnaK induced similar patterns
of immune response, so contaminants are not the only contributors to
stimulation of proPOase and TGase up-regulation. A novel mechanism for TGase
and proPOase increase in response to pathogens has been demonstrated.
Bizarrely, lower injections of both treatments induced stronger responses; this
inverted dose-response relationship points to an unknown mechanism of immune
system inhibition by DnaK. In conclusion, this study proposes the hypothesis
that bacterial derived HSP70 acts as an alarm signal, activating host TGase and
proPOase genes. I wonder how this response compares between organisms with only
innate immune system and those with both acquired and innate immune system; is
it stronger in the former because it is the main immune response? When pathogens
initially colonise a host, they are probably exposed to a sudden range of
stresses and therefore increase HSP production. This could be why the innate
immune system has evolved to respond to HSP, because high bacterial HSP levels
coincide with early infection.
Very interesting summary. Is interesting following on from my shrimp based paper which talks of the quasi-immune response (immune-like system) as a basis for allowing testing into vaccines.
ReplyDeleteClearly this paper backs up this possibility, detailing a possible method for inducing a response. Conclusion in which Bacterial produced HSP is very interesting, the immune-like system seeing it as a sign in which external pathogens may have colonised.
When you say that contamination of DnaK was less contaminated than in other experiments that have produced an immune response, does this mean that it was still contaminated? I imagine that is where the synDnaK comes in to control for contamination?
Immune system interactions with microbes is a great topic and one I will post about again. I bet invertebrate immune systems do have their own way of remembering pathogens.
DeleteThey say their DnaK endotoxin levels were less than the concentrations of pure endotoxins used in other studies to induce immune responses. But they were still contaminated, which is the reason for the uncontaminated control.