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FJ
EXPRESS SUMMARY ARTICLE The Full-length version of this article is also available, published online April 6, 2001 as doi:10.1096/fj.00-0709fje. |
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Laboratory of Molecular Endocrinology, CHUL Research Center and Department of Anatomy and Physiology, Laval University, Québec, Canada G1V 4G2
2Correspondence: Laboratory of Molecular Endocrinology, CHUL Research Center and Department of Anatomy and Physiology, Laval University, 2705, boul. Laurier, Québec, Canada G1V 4G2. E-mail: Serge.Rivest{at}crchul.ulaval.ca
SPECIFIC AIM
The complement system consists of a group of proteins that play essential roles in coordinating the host defense to infection. It can be activated by two primary pathways: the classical and alternative. The aim of this study was to determine the cellular distribution and the regulation of the genes encoding the proteins that are essential in guiding these pathways in the central nervous system (CNS) during innate immune recognition in mice.
PRINCIPAL FINDINGS
1. Constitutive expression of key members of the complement
proteins in the CNS
The results show a low to moderate C3 mRNA signal in few
non-neuronal structures under basal conditions that include the
epithelial lamina of the choroid plexus (chp), localized ependymal
cells lining the third ventricle, and the meninges of the ventral
medulla. In contrast, a robust C5 mRNA hybridization signal was found
in numerous populations of neuronal and non-neuronal cells, although
expression of the anaphylatoxin C5a receptor (C5aR) was low in the
brain of vehicle-administered mice. There was no constitutive
expression of the factor B and C3aR transcript in any regions of the
mouse brain.
2. Time-related induction in organs devoid of blood–brain barrier
(BBB)
The constitutive expression of C5 mRNA remained unaltered
during endotoxemia, whereas a strong and transient de novo expression
of the other members of the complement protein family was found in the
brain of mice that received a single systemic bolus of
lipopolysaccharide (LPS). Indeed, transcriptional activation C3 and
factor B genes occurred in the chp, the cerebral ependyma (C3 only),
leptomeninges and the circumventricular organs (CVOs), vascular organ
of the lamina terminalis, subfornical organ, median eminence, and the
area postrema (AP). The hybridization signal increased 3 h after
the injection, reached a maximal level from 6 to 12 h, and
vanished slowly to return to basal conditions 72 h after the
single LPS bolus.
3. Migratory-like pattern of LPS-induced C3aR gene expression
The results show no constitutive expression of the C3aR transcript
in any region of the mouse brain, but circulating LPS caused an
exquisite wave of C3aR-expressing cells from the leaky regions to
deeper parenchymal tissues. A positive signal was first detected along
the meninges, chp, and all the CVOs. These regions exhibited a positive
signal 3 h after the intraperitoneal (i.p.) LPS injection; the
intensity gradually increased to spread over the boundary of the CVOs,
meninges, and the walls of the ventricles 3 h later. Examples are
depicted in Fig. 1
, which shows C3aR-positive cells within the core of the AP 3 h
after a single i.p. LPS bolus (Fig. 1B
). Three hours later,
the number of C3aR-expressing cells increased to a maximum level within
the organ (Fig. 1C
). At that time, the C3aR hybridization
signal became visible along the AP boundary at the edge of the nucleus
of the solitary tract and gradually spread over the parenchymal brain
from 6 to 24 h post-LPS treatment (Fig. 1C
, D
, E
).
Positive C3aR-expressing cells were found across the entire brain
parenchyma 24 h after the injection, and the cerebral tissue
returned to basal levels 2 days later. Such migratory-like induction of
C3aR was also noted from all the other CVOs, the leptomeninges covering
the isocortex, and the ependymal lining wall of the cerebroventricular
system of endotoxin-challenged animals.
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4. Wave of C5aR-expressing cells from the cerebral capillaries to
the brain parenchyma
An interesting spreading effect of C5aR-expressing cells was
observed from the capillaries to their surrounding parenchymal regions.
Indeed, circulating LPS-sCD14 complex had the ability to target the
endothelial cells of the BBB and increased the expression of the
anaphylatoxin receptor C5aR. Few blood vessels became positive for the
C5aR transcript 1 h after the LPS injection (Fig. 2B
), whereas numerous microglial cells exhibited positive and
robust signal in the tissue adjacent to the capillary at 3 h (Fig. 2C
). The signal spread across the entire brain
parenchyma 6 h after the i.p. LPS bolus (Fig. 2D
) and
gradually returned to basal levels at 48 h.
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CONCLUSION
These data provide the evidence of a time-related regulation of different components of the complement system in the CNS and support the existence of an elegant innate immune response that takes place within specific populations of cells in the cerebral tissue during blood endotoxemia. The C3 and C5 are two major complement transcripts that are expressed in the CNS under basal conditions, although their patterns of expression were very different. Indeed, the mRNA encoding C3 was found in limited non-neuronal structures, whereas C5 mRNA was widely distributed across the brain parenchyma in both neuronal and non-neuronal elements. Such strong hybridization signal remained unchanged in response to circulating LPS, which differed from the gene encoding the C5aR that was up-regulated in the brain of LPS-challenged mice. A single bolus of endotoxin also caused a profound transcriptional activation of C3, C3aR, and factor B in numerous non-neuronal structures; the induction wave supported the concept of an integrated response of the complement system during endotoxemia. These genes are under a sophisticated transcriptional process in endothelial and myeloid-derived cells, because all induced transcripts returned to basal levels after the insult. The alternative pathway therefore exists in the brain, and this highly organized innate immune response may be essential for eliminating pathogens and orchestrating the inflammatory response to prevent the neuronal damage and restore the body homeostasis during systemic bacterial infection. Such systemic infection is censored by a group of supporting cells in regions devoid of BBB and the cerebral microvasculature in order to engage a time-related innate immune reaction necessary for preventing the neuronal damage, although it may have detrimental consequences when exaggerated.
As illustrated in Fig. 3
, the CVOs represent a route of entry for pathogens in the CNS, and C3
may tag the pathogen surface to increase the phagocytic activity of the
resident macrophages and microglia. The C3b (C3b-tagged pathogen) would
then promote the phagocytosis of the foreign pathogen by the binding of
C3b to its receptor, CR1 (CD35), expressed mainly on the surface of
leukocytes. De novo induction of the C3aR in the CNS may also be part
of the mechanisms that control the proinflammatory events to prepare
specific populations of cells to act immediately in the case of
pathogen invasion into the brain parenchyma via damaged or altered
blood vessels (see Fig. 3
). The integrity and physical characteristics
of the BBB are compromised during severe endotoxemia, allowing
diffusion of molecules that normally have no access to the cerebral
tissue and can be detrimental to the neuronal elements. Increasing
phagocytic activity of the CVO macrophages/microglia and adjacent
regions by the binding of a C3a fragment to its receptor, which are
both up-regulated in response to circulating LPS, may be essential to
prevent pathogen-induced neuronal damage.
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Complement factor B is a serine protease that plays a pivotal role in
the alternative pathway in the formation of the C3 convertase. As for
the C3aR, the mRNA encoding factor B was detectable only after a
systemic LPS challenge in different non-neuronal structures, namely,
the meninges, CVOs, chp, and along the endothelium of the brain
capillaries. The formation of C3 convertase at the surface of
C3b-tagged invading pathogens may then be possible by the presence of
factor B in the regions devoid of BBB (see Fig. 3
). Of particular
interest is the presence of the mRNAs encoding the factor B and C5aR
within scattered blood vessels that may either be more susceptible to
the infection and circulating pathogens or be the primary sentinels and
gatekeepers for the cerebral innate immune recognition. It is also
possible that uncontrolled expression of these molecules changes the
BBB properties featuring pathologies such as endotoxemia and cerebral
bacterial infections. Alteration of the BBB during severe endotoxemia
would open the way for immunological substances that have to be
recognized and processed. Activation of the microglial cells across the
CNS may rapidly eliminate this foreign material, although sustained
activity of these cells is not suitable.
There is accumulating evidence that chronic microglial reactivity is associated with neurodegenerative disorders. A better understanding of this innate immune response in the cerebral tissue may therefore lead us to the fundamental mechanisms underlying how the brain is capable of mounting inflammatory responses that either protect or contribute to damage neurons. The cerebral innate immunity is likely to be an essential player in the etiology of inflammatory CNS disorders resulting from infection as well as those assumed to have an immune etiology, such as multiple sclerosis. Since we are at the embryonic stage of the complement system in the brain, we speculate here that future studies will unravel unexpected findings supporting the concept that it directs and adapts the bilateral talk between the cerebral endothelium and microglia.
FOOTNOTES
1 To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.00-0709fje ; to
cite this article, use FASEB J. (April 6, 2001) 10.1096/fj.00-0709fje 
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