The role of cystatins in cells of the immune system

Article (PDF Available)inFEBS Letters 580(27):6295-301 · December 2006with16 Reads
DOI: 10.1016/j.febslet.2006.10.055 · Source: PubMed
The cystatins constitute a large group of evolutionary related proteins with diverse biological activities. Initially, they were characterized as inhibitors of lysosomal cysteine proteases - cathepsins. Cathepsins are involved in processing and presentation of antigens, as well as several pathological conditions such as inflammation and cancer. Recently, alternative functions of cystatins have been proposed: they also induce tumour necrosis factor and interleukin 10 synthesis and stimulate nitric oxide production. The aim of the present review was the analysis of data on cystatins from NCBI GEO database and the literature, and obtained in microarray and serial analysis of gene expression (SAGE) experiments. The expression of cystatins A, B, C, and F in macrophages, dendritic cells and natural killer cells of the immune system, during differentiation and activation is discussed.
The role of cystatins in cells of the immune system
a Kopitar-Jerala
Department of Biochemistry and Molecular Biology, Joz
ef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia
Received 16 August 2006; revised 22 October 2006; accepted 24 October 2006
Available online 3 November 2006
Edited by Masayuki Miyasaka
Abstract The cystatins constitute a large group of evolutionary
related proteins with diverse biological activities. Initially, they
were characterized as inhibitors of lysosomal cysteine proteases
cathepsins. Cathepsins are involved in processing and presenta-
tion of antigens, as well as several pathological conditions such
as inflammation and cancer. Recently, alternative functions of
cystatins have been proposed: they also induce tumour necrosis
factor and interleukin 10 synthesis and stimulate nitric oxide
production. The aim of the present review was the analysis of
data on cystatins from NCBI GEO database and the literature,
and obtained in microarray and serial analysis of gene expres-
sion (SAGE) experiments. The expression of cystatins A, B,
C, and F in macrophages, dendritic cells and natural killer cells
of the immune system, during differentiation and activation is
2006 Federation of European Biochemical Societies. Published
by Elsevier B.V. All rights reserved.
Keywords: Cystatin; Cathepsin; Macrophages; Dendritic cells;
Natural killer cells
1. Introduction
Cysteine cathepsins are long known to be responsible for
protein degradation in lysosomes. Recent studies show that
they are also involved in a number of other important cellular
processes such as antigen presentation [1], apoptosis, protein
processing [2], as well as several pathologies such as cancer
progression [3], inflammation [4] and neurodegeneration [5].
The role of lysosomal cathepsins in antigen presentation has
been reviewed recently [6]. Proteinase activity in these pro-
cesses is highly regulated at the level of protease expression,
by regulation of zymogen activation and by expression of
endogenous inhibitors. Natural inhibitors that inhibit cysteine
cathepsins include cystatins, thyrophins and also some of the
serpins [7–9].
The cystatins constitute a large group of evolutionary
related proteins acting as protease inhibitors of papain-like
cysteine proteases belonging to enzyme family C1 (see the
MEROPS database at, such as
cathepsins B, H, L, and S and legumain-related proteases of
the family C13 [10]. Type 1 cystatins, stefins (A and B), are
polypeptides of 98 amino acid residues which possess neither
disulfide bonds nor carbohydrate side chains and are located
mainly intracellularly. Type 2 cystatins C, D, E/M, F, S, SN,
and SA are characterized by two conserved disulfide bridges,
a larger size (120 residues) and the presence of a signal
peptide for extracellular targeting [11]. Type 3 cystatins, the
kininogens, are large (60–120 kDa) multifunctional plasma
proteins, containing three type 2 cystatin-like domains con-
taining a total of eight disulfide bridges. Although types 1
and 2 cystatins display considerable differences in amino acid
sequence, their tertiary structures are conserved and exhibit a
‘cystatin fold’ that is formed by five stranded anti-parallel b-
pleated sheet wrapped around a five-turn a-helix [12,13]. The
structure of human cystatin C in its dimeric form also shows
that each one of the two domains in the dimer adopt the typ-
ical monomeric ‘cystatin fold’ [14]. Some type 2 cystatins (C,
E/M, and F) are also able to inhibit mammalian legumain,
an asparaginyl endopeptidase (AEP), using a binding site dis-
tinct from the family C1 interaction site [15]. AEP has been
shown to be involved in class II major histocompatibility com-
plex (MHC) restricted antigen presentation [16] .
The present review focuses mostly on the expression of two
type 1 cystatins: stefins (cystatins) A and B and two type 2 cyst-
atins, cystatins C and F, in cells of the immune system upon
differentiation and activation. Two recently developed technol-
ogies, oligonucleotide or cDNA microarrays and serial analy-
sis of gene expression (SAGE), allow the determination of the
expression patterns of thousands of genes simultaneously
[17,18]. The gene expression omnibus (GEO) at the National
Center for Biotechnology Information (NCBI) is a large com-
pendium of gene expression data, addressing a wide range of
biological issues across many organisms [19] . The aim of the
present review is the identification of some of the most interest-
ing questions regarding cystatins in cells of the immune system
on the basis of recent data collected in the NCBI GEO and the
2. Stefin A (cystatin A) in follicular dendritic cells (FDC)
Stefin A (cystatin A) has been isolated from epidermis, poly-
morphonuclear granulocytes, liver, and spleen [20–23]. SAGE
Abbreviations: BCR, B cell receptor; DC, dendritic cells; FDC, follic-
ular dendritic cells; GC, germinal centres; SAGE, serial analysis of
gene expression; MHC, major histocompatibility complex; PKC,
protein kinase C; TNF-a, tumour necrosis factor alpha; IL-4, inter-
leukin-4; IL-10, interleukin-10; IFN-c, interferon-gamma; NK, natural
killer cells
Fax: +386 1 477 39 84.
E-mail address:
0014-5793/$32.00 Ó 2006 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
FEBS Letters 580 (2006) 6295–6301
studies showed that LPS stimulation decreased its synthesis in
monocytes [24]. Stefin A was also found in the follicular den-
dritic cells (FDC) in germinal centres (GC) of human tonsils
[25]. In contrast to antigen-presenting cells that present anti-
gens to T cells, FDC do not internalize, process and present
antigens in the context of major histocompatibility complex
class II (MHC II), but present intact antigen on their cell
surface [26,27]. GC form in lymphoid follicles of secondary
lymphoid organs and provide an essential microenvironment
for T cell-dependent humoral immune responses [28,29].
Within GC, antigen-specific B cells efficiently undergo clonal
expansion, isotype switching, somatic mutation, and affinity
maturation leading to the generation of plasma and memory
cells [30–32]. Only B-cells with the highest affinity B cell recep-
tor (BCR) bind to intact antigens on the surface of FDC and
receive survival signals from the FDC whereas low affinity
BCR B cells and self-reactive B cell clones are eliminated by
apoptosis [31]. Apoptosis in GC B-cells is mainly induced via
the death receptor pathway, by the rapid activation of cas-
pase-8 at the level of CD95 death-inducing signalling complex
(DISC) [33–35]. GC B cell apoptosis is dependent not only on
caspases but also endonucleases [36] and cathepsins [37]. Re-
cently, van Nierop et al. showed that apoptosis in human
GC B-cells involved lysosomal destabilization, which was con-
trolled by caspase-8 activity. CD40 ligation provided resistance
to lysosomal destabilization, as well as binding of high-affinity
B cells to FDC prevented lysosomal leakage and apoptosis in
GC B-cells [38]. Van Nierop et al. speculated that besides cas-
pase-8 inhibition there was an additional mechanism to pre-
vent lysosomal instability in adhering GC B-lymphocytes:
stefin A, which is expressed at high levels in FDC may play
a role in the prevention of apoptosis, as previously proposed
by van Eijk et al. [37,39].
3. Cystatins B and C in dendritic cells and macrophages
Cystatin C is the most potent inhibitor of cysteine proteases
such as cathepsins B, H, L, and S, with apparent inhibition
constants even below the nanomolar range [40]. Mature cysta-
tin C is synthesized as a preprotein with a 26 residue signal
peptide. Cystatin C is ubiquitously expressed in all tissues
and cell types, although mRNA levels vary several-fold be-
tween the tissues [41,42]. Oligomerization of cystatin C leads
to amyloid deposits in brain arteries at advanced age but this
pathological process is greatly accelerated in the mutant form
of cystatin C, responsible for hereditary cystatin C amyloid
angiopathy (HCCAA) [43]. Extracellular monomeric cystatin
C was found to be internalized by Chinese hamster ovary
(CHO) cells and trafficked into lysosomes where it dimerized
[44]. Cystatin C-deficient mice have essentially a normal phe-
notype. Cystatin C-deficient mice showed significantly reduced
growth of melanoma lung metastases when compared to wild-
type mice [45]. The reason for reduced growth of melanoma
lung metastases in cystatin C-deficient mice is unknown, but
could be a consequence of an early proteolytic event by a cys-
teine proteinase during the first hours after administration of
melanoma cells [45]. Cystatin C-deficient mice also showed
that cystatin C has a protective role in atherogenesis since cyst-
atin C-deficiency promotes atherosclerosis [46]. It was shown
that a glycosylated form of cystatin C is a necessary cofactor
for fibroblast growth factor 2 (FGF-2) induced mitogenic
activity on neural stem cells [47]. Cystatin C N-glycosylation
was necessary to induce neural stem cell proliferation. The pro-
tease inhibitory domain of cystatin C was not directly involved
in the process [47]. The unexpected consequences of cystatin
C deficiency on the spread of metastasis and atherosclerosis
could also be a consequence of alternative functions of cystatin
C, possibly as growth factor cofactor.
Dendritic cells (DC) are the professional antigen presenting
cells of the immune system. They are defined functionally by
their ability to take up antigens such as microorganisms, pro-
cess them into short antigenic peptides, load the peptides onto
major histocompatibility complex (MHC) molecules and then
present the resulting complexes at the cell surface. Immature
DC are located in the periphery of the body and they take
up and process antigens. Activated DC lose their capacity to
capture and process antigens. Instead they migrate to the sec-
ondary lymphoid organs and present antigen to T cells
Self peptides derived from secretory membrane proteins that
are synthesized by the antigen-presenting cells themselves bind
to MHC class II molecules tightly, but normally do not acti-
vate T cells. Cystatin C peptide (amino acids 40–55) has been
found as one of such self peptides bound to MHC class II mol-
ecules, indicating that it is endocytosed and cleaved with the
antigenic material and then bind to MHC class II molecules
[49]. Hashimoto et al. [50] observed that upon DC maturation
cystatin C transcripts were significantly downregulated (http:// The SAGE results of Hashim-
oto et al. were confirmed at the protein level by Zavasnik-Ber-
gant et al. who observed a large increase in intracellular
cystatin C during the differentiation of monocytes to immature
DC [51], Upon DC maturation, intracellular cystatin C levels
decreased and following prolonged incubation of mature DC
in the presence of TNF-a, cystatin C was secreted from DC.
It has been proposed that cystatin C plays a pivotal role in
the control of cleavage and removal of the MHC class II
invariant chain (Ii) by regulating the activity of cathepsin S,
and hence in the formation of MHC class II-peptide complexes
[52]. The work of El Sukkari et al. on DC isolated from cyst-
atin C-deficient mice showed that cystatin C is neither neces-
sary nor sufficient to control MHC class II expression and
antigen presentation in DC and that its expression differs be-
tween different DC subsets [53]. The absence of cystatin C
did not affect the expression, subcellular distribution, or for-
mation of peptide-loaded MHC class II complexes in any of
the DC types, nor the efficiency of presentation of exogenous
antigens [53]. Recent work by Kitamura et al. showed that
interleukin-6 (IL-6)-mediated signalling increased cathepsin S
activity, significantly reduced cystatin C expression and re-
duced the H2-DM and MHC class II ab dimer levels in DC
[54]. Overexpression of cystatin C in DC on the other hand sig-
nificantly suppressed IL-6-mediated enhancement of cathepsin
S activity and reduction of MHC class II ab dimer, Ii, and H2-
DM levels in DC. The authors concluded that the IL-6-medi-
ated alteration of the balance between cystatin C and cathepsin
S levels is important for the status of MHC class II ab dimer,
Ii, and H2-DM levels in DC. At least in the system described,
cystatin C may regulate cathepsin S activity in immature and
mature DC [54]. Murine spleen contains three major endo-
genous populations of DC. They are referred to as the CD8
, CD8
, and CD8
subsets [55,56]. CD8
DC are distinct from CD8
DC on the basis of a number of
criteria and primarily direct a Th2 response by activating T
6296 N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301
cells to secrete cytokines such as interleukin-4 (IL-4) [57,58].
DC produce IL-12 upon stimulation and induce a Th1
response [59]. Affymetrix microarray gene analysis was used
to determine gene expression patterns among murine DC sub-
sets: CD8
and CD8
cells were analyzed directly
after sorting and after 2 h cultivation (NCBI GEO GDS352).
Cystatin C was upregulated in cultured CD8
GEO GMS4772, GMS4773), an observation that is in agree-
ment with the study of El-Sukkari et al. [53]. DC also have
the capacity to take up, process and present exogenous anti-
gens in association with MHC class I molecules and this path-
way is termed cross-presentation [60–62]. CD8
DC have been
shown to be the principal DC subset involved in priming MHC
class I-restricted cytotoxic T cell immunity [63]. Since cystatin
C is expressed predominately in CD8
DCs, it is possible that it
has a role in this process.
Stefin B (cystatin B) is a type 1 cystatin that is distributed
rather uniformly among different tissues. In vitro stefin B binds
tightly to cathepsins H, L, and S, and less tightly to cathepsin
B [9]. Mutations in the gene encoding stefin B are responsible
for the primary defect in Unverricht-Lundborg disease
(EPM1) [64–66]. Stefin B-deficient mice display a phenotype
that is similar to the human disease with progressive ataxia
and myoclonic seizures [67]. The mice exhibit apoptosis of cer-
ebellar granullar cells and show increased expression of apop-
tosis and glial activation genes [68]. It was shown that removal
of cathepsin B from cystatin B-deficient mice greatly reduced
neuronal apoptosis, but did not rescue animals from ataxia
and seizure [69]. Thymocytes from stefin B-deficient mice ex-
erted a markedly increased response when they were exposed
to staurosporin, a protein kinase C (PKC) inhibitor compared
to thymocytes from wild-type mice [70]. We tested the possibil-
ity that stefin B interacts with the receptor for activated PKC
(RACK-1) in thymocytes and in this way interferes with PKC
signaling in the cells, but the interaction of RACK-1 with ste-
fin B was not confirmed. Preincubation of cells with E-64d did
not prevent apoptosis, indicating that staurosporin induces
apoptosis in a cathepsin-independent and caspase-dependent
manner. Brannvallet et al. reported that stefin B is localized
mainly in the nucleus of neural steam cells and in neurons,
while in glia cells it is also in the cytoplasm and in the lyso-
somes [71]. Hashimoto et al. showed that gene transcripts
of stefin B were significantly increased upon differentiation of
monocytes into macrophages [72]. However, upregulation of
the expression of the inhibitor upon differentiation of macro-
phages does not result in co-localization or interaction with
cathepsins L, S or B (Kopitar-Jerala, unpublished observa-
tions). The SAGE studies showed that treatment with LPS
causes upregulation of stefin B expression in human mono-
cytes, whereas cystatin C is not affected, indicating a possible
role of stefin B in innate immune response to bacterial infec-
tions [24].
Activated macrophages acquire antimicrobial activities
involving reactive oxygen species and reactive nitrogen metab-
olites. Chicken cystatin, cystatin C, and stefin B have been
implicated in nitric oxide (NO) production by interferon-c-
activated mouse peritoneal macrophages [73]. Mouse perito-
neal macrophages activated with interferon-gamma (IFN-c)
and then stimulated with IFN-c plus chicken cystatin gener-
ated increased amounts of NO in comparison with macro-
phages only activated with IFN-c [73]. The biological effect
of cystatins as NO synergistic inducers is not related to inhibi-
tion of a cysteine proteinase activity since E-64 did not induce
any increase in NO. Increased NO was due to increased induc-
ible NO synthase protein synthesis. Further studies of Verdot
et al. suggested that chicken cystatin stimulated the release of
TNF-a and interleukin-10 (IL-10) by IFN- c-activated murine
peritoneal macrophages [74]. This observation could be of bio-
logical importance as cystatin concentrations necessary to
upregulate TNF-a, IL-10 and NO synthesis are in the physio-
logical range, as found in human body fluids. The cystatin C-
mediated release of TNF-a is probably responsible for the
increase in NO production by IFN-c-activated murine perito-
neal macrophages. The findings by Verdot et al. [74] point at
a new relationship between cystatins, cytokines, inflammation
and immune responses.
In vitro experiments in cell culture models described above
were confirmed by experiments in Leishmania donovani in-
fected mice [75]. L. donovani, the etiological agent for the
severe visceral form of leishmaniasis, multiplies in the phago-
lysosomes of macrophages of the infected host. Treatment of
L. donovani-infected murine peritoneal macrophages with a
combination of chicken cystatin and IFN-c induced increased
production of NO and did overcome the inhibition of NO
synthesis driven by L. donovani parasites. Mice treated with
chicken cystatin and IFN-c showed reduced splenomegaly, a
lowered parasite burden in the spleen and increased produc-
tion of NO [75]. The infected mice treated with chicken cysta-
tin and IFN-c were cured by the induction of NO that killed
the parasite and the switched CD4
T cell-mediated immune
responses from disease-promoting Th2 cells to the protective
Th1 response shown by the increased production of IL-12
and decreased production of IL-4.
4. Cystatin F in NK cells
Cystatin F is expressed in a variety of tissues. Expression is
particularly high in the cells and tissues of the immune system:
thymus and spleen, monocytes, DC, T-cells and NK cells [76].
Mature cystatin F is composed of 126 amino acid residues. It is
synthesized as a preprotein with a 19 residue signal peptide and
possesses a unique extension of six amino acids at its N-termi-
nus. In addition to the two disulfide bridges common to all
type 2 cystatins, mature cystatin F has two cysteine residues
that, form an interinolecular disulfide bridge, as revealed in
the crystal structure of the cystatin F dimer [77]. Cappello
et al. observred that in U937 cells cystatin F was secreted as
a disulfide bridge-linked dimer [78]. Cystatin F dimer is inac-
tive as an inhibitor of papain like cathepsins and can be
activated by chemical reduction [79]. As compared to other
cystatins, the protein exhibits a distinct inhibitory profile. It
binds tightly to cathepsins F, K, L, and V, less tightly to
cathepsins S and H, and does not inhibit cathepsins B, C,
and X [79]. Cystatin F can inhibit AEP, but with lower affinity
as cystatin E/M and C [80]. The recently elucidated crystal
structure of cystatin F revealed that two N-linked glycosyla-
tion sites of cystatin F may modulate its inhibitory properties,
in particular its reduced affinity towards AEP as compared to
other cystatins [77].
Cystatin F has also been shown to be strongly upregulated
in LPS-stimulated monocyte-derived DC [50]. In the U937
premyeloid cell line, both differentiation towards a granulo-
cytic pathway by all-trans-retinoic acid (ATRA) or towards
N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301 6297
a monocytic pathway by stimulation with phorbol ester (TPA)
resulted in marked downregulation of cystatin F expression
In U937 cells, cystatin F has been found to be secreted and
localized intracellularly in lysosome-like granules [81]. Cap-
pello et al. found it in lysosomes in transfected HeLa cells,
but not in U937 cells. Sorting of cystatin F to lysosomes was
greatly enhanced when its C-terminal end was extended by
few amino acids [78]. The authors concluded that under partic-
ular conditions, cystatin F can be sorted to the endocytic path-
way, but its unusual inhibitory function is mainly performed
extracellularly and probably controlled through dimerization
[78]. Langerholc et al. showed that in U937 cells, cystatin F
was co-localized with lysosomal markers LAMP-2 and CD68
and when subcellular localization of cystatin F was compared
to that of cathepsins, cystatin F was found to be co-localized
with cathepsins X and H, but not with cathepsins L, B, C
[79]. Further investigations on cystatin F localization, possibly
in other cells and in cystatin F-deficient mice, will be necessary
to elucidate its biological function (see Fig. 1).
Gene expression analysis of human NK cells and CD8
lymphocytes revealed that transcripts of cystatin F were sign-
ificantly upregulated in NK cells when compared to CD8
lymphocytes ( [82].
NK cells are innate immune lymphocytes that mediate two
major functions: recognition and lysis of cancer cells and
virus-infected cells and production of immunoregulatory cyto-
kines [83,84]. The activation of NK cells is controlled by com-
plex interactions between activating and inhibitory receptor
signals and can be modulated by cytokines [85]. Human NK
cells comprise approximately 15% of peripheral blood lympho-
cytes and the majority of human NK cells are CD56
whereas a minority are CD56
and CD16
[84]. The
function of CD56
NK cells is different from that of
NK cells. CD56
NK cells can produce cytokines
more abundantly, consistent with their functional role as an in-
nate immunoregulator [86]. In contrast, CD56
cells seem to be skewed toward homing to inflammation sites
and promoting immune responses, in addition to induction
of cytotoxicity [86].
Hanna and coworkers reported that cystatin F tran-
scripts were more abundant in CD56
NK cells and
in vitro activated CD56
NK cells than in
NK cells [87] (NCBI GEO GSM26200-5).
Although the microarray data do not give us any information
about inhibitory activity of cystatin F and have to be inter-
preted with caution, it is tempting to speculate that cystatin
F plays a specific role in the function of NK (CD56
5. Conclusions
Several structural and kinetic studies have given us insight
into interactions of cystatins and cysteine cathepsins in vitro
but in vivo very few interactions have been found. The present
review aims to identify the most interesting questions rather
than providing the definitive answers. The central question
remains what the targets of cystatins are that are differentially
regulated in cells of the immune system. Is it possible that cyst-
atins have different roles in different tissues like serpins? For
example, tPA is not just a ‘plasminogen activator’; it is now
Fig. 1. Stefin A (cystatin A) is expressed at high levels in FDC and may play a role in the prevention of apoptosis in GC B cells as proposed by van
Eijk et al. [37,39]. Several signalling molecules are involved in FDC-GC B cell contacts: intercellular adhesion molecule-1 (ICAM-1) and vascular cell
adhesion molecule-1 (VCAM-1) enhance cell-cell contact; B-cell-activating factor of the tumour necrosis factor family (BAFF/BLys) prevents
apoptosis of GC B cells and interleukin-15 (IL-15) stimulate GC B-cell proliferation [28].
6298 N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301
widely appreciated for its role in the central nervous system
[88,89]. Although it can act on its classical substrate, plasmin-
ogen, it also associates with other targets, and in some cases
can even act like a cytokine to activate microglial cells without
engaging its catalytic properties [90].
The real challenge that lies in front of us is to discover pro-
teinases (and possibly some other proteins) which interact with
cystatins that are differentially upregulated in cells of the im-
mune system.
Acknowledgements: This work was supported by the Ministry of High
Education, Science and Technology of the Republic of Slovenia. Prof.
R.H. Pain is gratefully acknowledged for critically reading the manu-
script, giving useful comments and editing English. I also thank Prof.
B. Turk and Prof. V. Turk for reading the manuscript and giving use-
ful suggestions.
[1] Honey, K. and Rudensky, A.Y. (2003) Lysosomal cysteine
proteases regulate antigen presentation. Nat. Rev. Immunol. 3,
[2] Turk, V., Turk, B. and Turk, D. (2001) Lysosomal cysteine
proteases: facts and opportunities. EMBO J. 20, 4629–4633.
[3] Kos, J. and Lah, T.T. (1998) Cysteine proteinases and their
endogenous inhibitors: target proteins for prognosis, diagnosis
and therapy in cancer (review). Oncol. Rep. 5, 1349–1361.
[4] Lang, A., Horler, D. and Baici, A. (2000) The relative importance
of cysteine peptidases in osteoarthritis. J. Rheumatol. 27, 1970–
[5] Nixon, R.A., Cataldo, A.M. and Mathews, P.M. (2000) The
endosomal–lysosomal system of neurons in Alzheimer’s disease
pathogenesis: a review. Neurochem. Res. 25, 1161–1172.
[6] Hsing, L.C. and Rudensky, A.Y. (2005) The lysosomal cysteine
proteases in MHC class II antigen presentation. Immunol. Rev.
207, 229–241.
[7] Lenarcic, B. and Bevec, T. (1998) Thyropins new structurally
related proteinase inhibitors. Biol. Chem. 379, 105–111.
[8] Liu, N., Raja, S.M., Zazzeroni, F., Metkar, S.S., Shah, R., Zhang,
M., Wang, Y., Bromme, D., Russin, W.A., Lee, J.C., Peter, M.E.,
Froelich, C.J., Franzoso, G. and Ashton-Rickardt, P.G. (2003)
NF-kappaB protects from the lysosomal pathway of cell death.
EMBO J. 22, 5313–5322.
[9] Turk, B., Turk, V. and Turk, D. (1997) Structural and functional
aspects of papain-like cysteine proteinases and their protein
inhibitors. Biol. Chem. 378, 141–150.
[10] Rawlings, N.D., Morton, F.R. and Barrett, A.J. (2006) MER-
OPS: the peptidase database. Nucleic Acids Res. 34, D270–D272.
[11] Rawlings, N.D., Tolle, D.P. and Barrett, A.J. (2004) Evolutionary
families of peptidase inhibitors. Biochem. J. 378, 705–716.
[12] Bode, W., Engh, R., Musil, D., Thiele, U., Huber, R., Karshikov,
A., Brzin, J., Kos, J. and Turk, V. (1988) The 2.0 A
X-ray crystal
structure of chicken egg white cystatin and its possible mode of
interaction with cysteine proteinases. EMBO J. 7, 2593–2599.
[13] Stubbs, M.T., Laber, B., Bode, W., Huber, R., Jerala, R.,
Lenarcic, B. and Turk, V. (1990) The refined 2.4 A
X-ray crystal
structure of recombinant human stefin B in complex with the
cysteine proteinase papain: a novel type of proteinase inhibitor
interaction. EMBO J. 9, 1939–1947.
[14] Janowski, R., Kozak, M., Jankowska, E., Grzonka, Z., Grubb,
A., Abrahamson, M. and Jaskolski, M. (2001) Human cystatin C,
an amyloidogenic protein, dimerizes through three-dimensional
domain swapping. Nat. Struct. Biol. 8, 316–320.
[15] Alvarez-Fernandez, M., Barrett, A.J., Gerhartz, B., Dando, P.M.,
Ni, J. and Abrahamson, M. (1999) Inhibition of mammalian
legumain by some cystatins is due to a novel second reactive site.
J. Biol. Chem. 274, 19195–19203.
[16] Manoury, B., Hewitt, E.W., Morrice, N., Dando, P.M., Barrett,
A.J. and Watts, C. (1998) An asparaginyl endopeptidase processes
a microbial antigen for class II MHC presentation. Nature 396,
[17] Chee, M., Yang, R., Hubbell, E., Berno, A., Huang, X.C., Stern,
D., Winkler, J., Lockhart, D.J., Morris, M.S. and Fodor, S.P.
(1996) Accessing genetic information with high-density DNA
arrays. Science 274, 610–614.
[18] Velculescu, V.E., Zhang, L., Vogelstein, B. and Kinzler, K.W.
(1995) Serial analysis of gene expression. Science 270, 484–487.
[19] Barrett, T., Suzek, T.O., Troup, D.B., Wilhite, S.E., Ngau, W.C.,
Ledoux, P., Rudnev, D., Lash, A.E., Fujibuchi, W. and Edgar, R.
(2005) NCBI GEO: mining millions of expression profiles-
database and tools. Nucleic Acids Res. 33, D562–D566.
[20] Brzin, J., Kopitar, M., Locnikar, P. and Turk, V. (1982) An
endogenous inhibitor of cysteine and serine proteinases from
spleen. FEBS Lett. 138, 193–197.
[21] Brzin, J., Kopitar, M., Turk, V. and Machleidt, W. (1983) Protein
inhibitors of cysteine proteinases. I. Isolation and characterization
of stefin, a cytosolic protein inhibitor of cysteine proteinases from
human polymorphonuclear granulocytes. Hoppe Seylers Z.
Physiol Chem. 364, 1475–1480.
[22] Green, G.D., Kembhavi, A.A., Davies, M.E. and Barrett, A.J.
(1984) Cystatin-like cysteine proteinase inhibitors from human
liver. Biochem. J. 218, 939–946.
[23] Jarvinen, M. (1978) Purification and some characteristics of the
human epidermal SH-protease inhibitor. J. Invest Dermatol. 71,
[24] Suzuki, T., Hashimoto, S., Toyoda, N., Nagai, S., Yamazaki, N.,
Dong, H.Y., Sakai, J., Yamashita, T., Nukiwa, T. and Matsu-
shima, K. (2000) Comprehensive gene expression profile of LPS-
stimulated human monocytes by SAGE. Blood 96, 2584–2591.
[25] Rinne, A., Dorn, A., Jarvinen, M., Alavaikko, M., Jokinen, K.
and Hopsu-Havu, V.K. (1986) Immunoelectron microscopical
location of the acid cysteine proteinase inhibitor in the lymphatic
tissue of the tonsils. Acta Histochem. 79, 137–145.
[26] Burton, G.F., Conrad, D.H., Szakal, A.K. and Tew, J.G. (1993)
Follicular dendritic cells and B cell costimulation. J. Immunol.
150, 31–38.
[27] Fu, Y.X. and Chaplin, D.D. (1999) Development and maturation
of secondary lymphoid tissues. Annu. Rev. Immunol. 17, 399–
[28] Park, C.S. and Choi, Y.S. (2005) How do follicular dendritic cells
interact intimately with B cells in the germinal centre? Immuno-
logy 114, 2–10.
[29] Tew, J.G., Wu, J., Qin, D., Helm, S., Burton, G.F. and Szakal,
A.K. (1997) Follicular dendritic cells and presentation of antigen
and costimulatory signals to B cells. Immunol. Rev. 156, 39–52.
[30] Liu, Y.J., Joshua, D.E., Williams, G.T., Smith, C.A., Gordon, J.
and MacLennan, I.C. (1989) Mechanism of antigen-driven
selection in germinal centres. Nature 342, 929–931.
[31] Liu, Y.J., Arpin, C., de Bouteiller, O., Guret, C., Banchereau, J.,
Martinez-Valdez, H. and Lebecque, S. (1996) Sequential trigger-
ing of apoptosis, somatic mutation and isotype switch during
germinal center development. Semin. Immunol. 8, 169–177.
[32] MacLennan, J.C. (1994) Germinal centers. Annu. Rev. Immunol.
12, 117–139.
[33] Hennino, A., Berard, M., Krammer, P.H. and Defrance, T. (2001)
FLICE-inhibitory protein is a key regulator of germinal center B
cell apoptosis. J. Exp. Med. 193, 447–458.
[34] van Eijk, M., Medema, J.P. and de Groot, C. (2001) Cutting edge:
cellular Fas-associated death domain-like IL-1-converting
enzyme-inhibitory protein protects germinal center B cells from
apoptosis during germinal center reactions. J. Immunol. 166,
[35] van Eijk, M., Defrance, T., Hennino, A. and de Groot, C. (2001)
Death-receptor contribution to the germinal-center reaction.
Trends Immunol. 22, 677–682.
[36] Lindhout, E., Lakeman, A. and de Groot, C. (1995) Follicular
dendritic cells inhibit apoptosis in human B lymphocytes by a
rapid and irreversible blockade of preexisting endonuclease. J.
Exp. Med. 181, 1985–1995.
[37] van Eijk, M. and de Groot, C. (1999) Germinal center B cell
apoptosis requires both caspase and cathepsin activity. J. Immu-
nol. 163, 2478–2482.
[38] van Nierop, K., Muller, F.J., Stap, J., Van Noorden, C.J., van
Eijk, M., and de Groot, C. (2006). Lysosomal destabilization
contributes to apoptosis of germinal center B-lymphocytes. J.
Histochem. Cytochem. doi:10.1369/jhc.6A6967.2006
N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301 6299
[39] van Eijk, M., Van Noorden, C.J. and de Groot, C. (2003)
Proteinases and their inhibitors in the immune system. Int. Rev.
Cytol. 222, 197–236.
[40] Lindahl, P., Abrahamson, M. and Bjork, I. (1992) Interaction of
recombinant human cystatin C with the cysteine proteinases
papain and actinidin. Biochem. J. 281, 49–55.
[41] Abrahamson, M., Barrett, A.J., Salvesen, G. and Grubb, A.
(1986) Isolation of six cysteine proteinase inhibitors from human
urine. Their physicochemical and enzyme kinetic properties and
concentrations in biological fluids. J. Biol. Chem. 261, 11282–
[42] Abrahamson, M., Olafsson, I., Palsdottir, A., Ulvsback, M.,
Lundwall, A., Jensson, O. and Grubb, A. (1990) Structure and
expression of the human cystatin C gene. Biochem. J. 268, 287–
[43] Abrahamson, M., Jonsdottir, S., Olafsson, I., Jensson, O. and
Grubb, A. (1992) Hereditary cystatin C amyloid angiopathy:
identification of the disease-causing mutation and specific diag-
nosis by polymerase chain reaction based analysis. Hum. Genet.
89, 377–380.
[44] Merz, G.S., Benedikz, E., Schwenk, V., Johansen, T.E., Vogel,
L.K., Rushbrook, J.L. and Wisniewski, H.M. (1997) Human
cystatin C forms an inactive dimer during intracellular trafficking
in transfected CHO cells. J. Cell Physiol. 173, 423–432.
[45] Huh, C.G., Hakansson, K., Nathanson, C.M., Thorgeirsson,
U.P., Jonsson, N., Grubb, A., Abrahamson, M. and Karlsson, S.
(1999) Decreased metastatic spread in mice homozygous for a null
allele of the cystatin C protease inhibitor gene. Mol. Pathol. 52,
[46] Bengtsson, E., To, F., Hakansson, K., Grubb, A., Branen, L.,
Nilsson, I. and Jovinge, S. (2005) Lack of the cysteine protease
inhibitor cystatin C promotes atherosclerosis in apolipoprotein
E-deficient mice. Arterioscler. Thromb. Vase. Biol. 25, 2151–2156.
[47] Taupin, P., Ray, J., Fischer, W.H., Suhr, S.T., Hakansson, K.,
Grubb, A. and Gage, F.H. (2000) FGF-2-responsive neural stem
cell proliferation requires CCg, a novel autocrine/paracrine
cofactor. Neuron 28, 385–397.
[48] Mellman, I. and Steinman, R.M. (2001) Dendritic cells: special-
ized and regulated antigen processing machines. Cell 106, 255–
[49] Hunt, D.F., Michel, H., Dickinson, T.A., Shabanowitz, J., Cox,
A.L., Sakaguchi, K., Appella, E., Grey, H.M. and Sette, A. (1992)
Peptides presented to the immune system by the murine class II
major histocompatibility complex molecule I-Ad. Science 256,
[50] Hashimoto, S., Suzuki, T., Dong, H.Y., Nagai, S., Yamazaki, N.
and Matsushima, K. (1999) Serial analysis of gene expression in
human monocyte-derived dendritic cells. Blood 94, 845–852.
[51] Zavasnik-Bergant, T., Repnik, U., Schweiger, A., Romih, R.,
Jeras, M., Turk, V. and Kos, J. (2005) Differentiation- and
maturation-dependent content, localization, and secretion of
cystatin C in human dendritic cells. J. Leukoc. Biol. 78, 122–134.
[52] Pierre, P. and Mellman, I. (1998) Developmental regulation of
invariant chain proteolysis controls MHC class II trafficking in
mouse dendritic cells. Cell 93, 1135–1145.
[53] El Sukkari, D., Wilson, N.S., Hakansson, K., Steptoe, R.J.,
Grubb, A., Shortman, K. and Villadangos, J.A. (2003) The
protease inhibitor cystatin C is differentially expressed among
dendritic cell populations, but does not control antigen presen-
tation. J. Immunol. 171, 5003–5011.
[54] Kitamura, H., Kamon, H., Sawa, S., Park, S.J., Katunuma, N.,
Ishihara, K., Murakami, M. and Hirano, T. (2005) IL-6-STAT3
controls intracellular MHC class II alphabeta dimer level through
cathepsin S activity in dendritic cells. Immunity 23, 491–502.
[55] Edwards, A.D., Chaussabel, D., Tomlinson, S., Schulz, O., Sher,
A. and Reis e Sousa, C. (2003) Relationships among murine
CD11c(high) dendritic cell subsets as revealed by baseline gene
expression patterns. J. Immunol. 171, 47–60.
[56] Vremec, D., Pooley, J., Hochrein, H., Wu, L. and Shortman, K.
(2000) CD4 and CDS expression by dendritic cell subtypes in
mouse thymus and spleen. J. Immunol. 164, 2978–2986.
[57] Pulendran, B., Smith, J.L., Caspary, G., Brasel, K., Pettit, D.,
Maraskovsky, E. and Maliszewski, C.R. (1999) Distinct dendritic
cell subsets differentially regulate the class of immune response
in vivo. Proc. Natl Acad. Sci. USA 96, 1036–1041.
[58] Maldonado-Lopez, R., De Smedt, T., Michel, P., Godfroid, J.,
Pajak, B., Heirman, C., Thielemans, K., Leo, O., Urbain, J. and
Moser, M. (1999) CD8alpha+ and CD8alpha-subclasses of
dendritic cells direct the development of distinct T helper cells
in vivo. J. Exp. Med. 189, 587–592.
[59] Hochrein, H., Shortman, K., Vremec, D., Scott, B., Hertzog, P.
and O’Keeffe, M. (2001) Differential production of IL-12, IFN-
alpha, and IFN-gamma by mouse dendritic cell subsets. J.
Immunol. 166, 5448–5455.
[60] den Haan, J.M., Lehar, S.M. and Bevan, M.J. (2000) CD8(+) but
not CD8() dendritic cells cross-prime cytotoxic T cells in vivo. J.
Exp. Med. 192, 1685–1696.
[61] den Haan, J.M. and Bevan, M.J. (2001) Antigen presentation to
CD8+T cells: cross-priming in infectious diseases. Curr. Opin.
Immunol. 13, 437–441.
[62] Heath, W.R., Belz, G.T., Behrens, G.M., Smith, C.M., Forehan,
S.P., Parish, J.A., Davey, G.M., Wilson, N.S., Carbone, F.R. and
Villadangos, J.A. (2004) Cross-presentation, dendritic cell subsets,
and the generation of immunity to cellular antigens. Immunol.
Rev. 199, 9–26.
[63] Belz, G.T., Shortman, K., Bevan, M.J. and Heath, W.R. (2005)
CD8alpha+ dendritic cells selectively present MHC class I-
restricted noncytolytic viral and intracellular bacterial antigens
in vivo. J. Immunol. 175, 196–200.
[64] Lalioti, M.D., Mirotsou, M., Buresi, C., Peitsch, M.C., Rossier,
C., Ouazzani, R., Baldy-Moulinier, M., Bottani, A., Malafosse,
A. and Antonarakis, S.E. (1997) Identification of mutations in
cystatin B, the gene responsible for the Unverricht-Lundborg type
of progressive myoclonus epilepsy (EPM1). Am. J. Hum. Genet.
60, 342–351.
[65] Lalioti, M.D., Scott, H.S., Buresi, C., Rossier, C., Bottani, A.,
Morris, M.A., Malafosse, A. and Antonarakis, S.E. (1997)
Dodecamer repeat expansion in cystatin B gene in progressive
myoclonus epilepsy. Nature 386, 847–851.
[66] Pennacchio, L.A., Lehesjoki, A.E., Stone, N.E., Willour, V.L.,
Virtaneva, K., Miao, J., D’Amato, E., Ramirez, L., Faham, M.,
Koskiniemi, M., Warrington, J.A., Norio, R., de la Chapelle, A.,
Cox, D.R. and Myers, R.M. (1996) Mutations in the gene
encoding cystatin B in progressive myoclonus epilepsy (EPM1).
Science 271, 1731–1734.
[67] Pennacchio, L.A., Bouley, D.M., Higgins, K.M., Scott, M.P.,
Noebels, J.L. and Myers, R.M. (1998) Progressive ataxia,
myoclonic epilepsy and cerebellar apoptosis in cystatin B-deficient
mice. Nat. Genet. 20, 251–258.
[68] Lieuallen, K., Pennacchio, L.A., Park, M., Myers, R.M. and
Lennon, G.G. (2001) Cystatin B-deficient mice have increased
expression of apoptosis and glial activation genes. Hum. Mot
Genet. 10, 1867–1871.
[69] Houseweart, M.K., Pennacchio, L.A., Vilaythong, A., Peters, C.,
Noebels, J.L. and Myers, R.M. (2003) Cathepsin B but not
cathepsins L or S contributes to the pathogenesis of Unverricht-
Lundborg progressive myoclonus epilepsy (EPM1). J. Neurobiol.
56, 315–327.
[70] Kopitar-Jerala, N., Schweiger, A., Myers, R.M., Turk, V. and
Turk, B. (2005) Sensitization of stefin B-deficient thymocytes
towards staurosporin-induced apoptosis is independent of cys-
teine cathepsins. FEBS Lett. 579, 2149–2155.
[71] Brannvall, K., Hjelm, H., Korhonen, L., Lahtinen, U., Lehesjoki,
A.E. and Lindholm, D. (2003) Cystatin-B is expressed by neural
stem cells and by differentiated neurons and astrocytes. Biochem.
Biophys. Res. Commun. 308, 369–374.
[72] Hashimoto, S., Suzuki, T., Dong, H.Y., Yamazaki, N. and
Matsushima, K. (1999) Serial analysis of gene expression in
human monocytes and macrophages. Blood 94, 837–844.
[73] Verdot, L., Lalmanach, G., Vercruysse, V., Hartmann, S., Lucius,
R., Hoebeke, J., Gauthier, F. and Vray, B. (1996) Cystatins up-
regulate nitric oxide release from interferon-gamma-activated
mouse peritoneal macrophages. J. Biol. Chem. 271, 28077–28081.
[74] Verdot, L., Lalmanach, G., Vercruysse, V., Hoebeke, J., Gau-
thier, F. and Vray, B. (1999) Chicken cystatin stimulates nitric
oxide release from interferon-gamma-activated mouse peritoneal
macrophages via cytokine synthesis. Eur. J. Biochem. 266, 1111–
[75] Das, L., Datta, J.N.L., Bandyopadhyay, S. and Das, P.K. (2001)
Successful therapy of lethal murine visceral leishmaniasis with
6300 N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301
cystatin involves up-regulation of nitric oxide and a favorable T
cell response. J. Immunol. 166, 4020–4028.
[76] Ni, J., Fernandez, M.A., Danielsson, L., Chillakuru, R.A.,
Zhang, J., Grubb, A., Su, J., Gentz, R. and Abrahamson, M.
(1998) Cystatin F is a glycosylated human low molecular weight
cysteine proteinase inhibitor. J. Biol. Chem. 273, 24797–24804.
[77] Schuttelkopf, A.W., Hamilton, G., Watts, C. and van Aalten,
D.M. (2006) Structural basis of reduction-dependent activation of
human Cystatin F. J. Biol. Chem. 281, 16570–16575.
[78] Cappello, F., Gatti, E., Camossetto, V., David, A., Lelouard, H.
and Pierre, P. (2004) Cystatin F is secreted, but artificial
modification of its C-terminus can induce its endocytic targeting.
Exp. Cell Res. 297, 607–618.
[79] Langerholc, T., Zavasnik-Bergant, V., Turk, B., Turk, V.,
Abrahamson, M. and Kos, J. (2005) Inhibitory properties of
cystatin F and its localization in U937 promonocyte cells. FEBS J.
272, 1535–1545.
[80] Alvarez-Fernandez, M., Barrett, A.J., Gerhartz, B., Dando, P.M.,
Ni, J. and Abrahamson, M. (1999) Inhibition of mammalian
legumain by some cystatins is due to a novel second reactive site.
J. Biol. Chem. 274, 19195–19203.
[81] Nathanson, C.M., Wasselius, J., Wallin, H. and Abrahamson, M.
(2002) Regulated expression and intracellular localization of
cystatin F in human U937 cells. Eur. J. Biochem. 269, 5502–5511.
[82] Obata-Onai, A., Hashimoto, S., Onai, N., Kurachi, M., Nagai, S.,
Shizuno, K., Nagahata, T. and Matsushima, K. (2002) Compre-
hensive gene expression analysis of human NK cells and CD8(+)
T lymphocytes. Int. Immunol. 14, 1085–1098.
[83] Cooper, M.A., Fehniger, T.A. and Caligiuri, M.A. (2001) The
biology of human natural killer-cell subsets. Trends Immunol. 22,
[84] Robertson, M.J. and Ritz, J. (1990) Biology and clinical relevance
of human natural killer cells. Blood 76, 2421–2438.
[85] Lanier, L.L. (2001) Face off-the interplay between activating
and inhibitory immune receptors. Curr. Opin. Immunol. 13, 326–
[86] Cooper, M.A., Fehniger, T.A., Turner, S.C., Chen, K.S., Gha-
heri, B.A., Ghayur, T., Carson, W.E. and Caligiuri, M.A. (2001)
Human natural killer cells: a unique innate immunoregulatory
role for the CD56(bright) subset. Blood 97, 3146–3151.
[87] Hanna, J., Bechtel, P., Zhai, Y., Youssef, F., McLachlan, K. and
Mandelboim, O. (2004) Novel insights on human NK cells,
immunological modalities revealed by gene expression profiling.
J. Immunol. 173, 6547–6563.
[88] Nicole, O., Docagne, F., Ali, C., Margaill, J., Carmeliet, P.,
MacKenzie, E.T., Vivien, D. and Buisson, A. (2001) The
proteolytic activity of tissue-plasminogen activator enhances
NMDA receptor-mediated signaling. Nat. Med. 7, 59–64.
[89] Tsirka, S.E. (2002) Tissue plasminogen activator as a modulator
of neuronal survival and function. Biochem. Soc. Trans. 30, 222–
[90] Rogove, A.D., Siao, C., Keyt, B., Strickland, S. and Tsirka, S.E.
(1999) Activation of microglia reveals a non-proteolytic cytokine
function for tissue plasminogen activator in the central nervous
system. J. Cell Sci. 112, 4007–4016.
N. Kopitar-Jerala / FEBS Letters 580 (2006) 6295–6301 6301
    • "Many of these proteins (actin, cystatin, desmoplakin, filaggrin, hornerin, calgranulin and zinc-alpha-2-glycoprotein) are localised in the epithelial lining and are involved in protein binding, cell adhesion and cytoskeletal structure, and inflammatory processes @BULLET One protease inhibitor, cystatin A, was present with a particularly high abundance (3%) of total EBC protein in the Muccilli profile of healthy subjects [44]. Cystatin A induces synthesis of TNF-α and IL-10 [114] and is closely associated with inflammation and cancer [115] @BULLET Another molecule that has drawn attention since its description in EBC by Carpagnano et al [116] is endothelin, a pro-inflammatory, pro-fibrotic, broncho-and vasoconstrictive peptide. Other similar proteomic research undertaken in EBC by Carpagnano et al has also investigated the potential of leptin [94], matrix metalloproteinase-9 [117], cycloxygenase II and survivin [118] for diagnosing NSCLC Cigarette smoking induces an inflammatory response in the airways that may play a key role in the pathogenesis of LC, therefore it is imperative that research is done to characterise the effect of smoking on total protein, as well as individual protein biomarkers. "
    [Show abstract] [Hide abstract] ABSTRACT: Lung cancer is a leading cause of cancer-related deaths worldwide, and is considered one of the most aggressive human cancers, with a 5 year overall survival of 10-15%. Early diagnosis of lung cancer is ideal; however, it is still uncertain as to what technique will prove successful in the systematic screening of high-risk populations, with the strongest evidence currently supporting low dose computed tomography (LDCT). Analysis of exhaled breath condensate (EBC) has recently been proposed as an alternative low risk and non-invasive screening method to investigate early-stage neoplastic processes in the airways. However, there still remains a relative paucity of lung cancer research involving EBC, particularly in the measurement of lung proteins that are centrally linked to pathogenesis. Considering the ease and safety associated with EBC collection, and advances in the area of mass spectrometry based profiling, this technology has potential for use in screening for the early diagnosis of lung cancer. This review will examine proteomics as a method of detecting markers of neoplasia in patient EBC with a particular emphasis on LC, as well as discussing methodological challenges involving in proteomic analysis of EBC specimens.
    Full-text · Article · Jul 2016
    • "Type-3 cystatins are high molecular weight (60–120 kDa) proteins and have three repeated type 2-like cystatin domains (Salvesen et al., 1986). Cystatins in immune cells have been reported to participate in the release of NO, phagocytosis and expression of cytokines (Kopitar-Jerala, 2006; Magister and Kos, 2013; Maher et al., 2014a). Stefin B belongs to the type one cystatins and is located in the cytosol, mitochondria and nucleus (Ceru et al., 2010; Maher et al., 2014a). "
    [Show abstract] [Hide abstract] ABSTRACT: Recently several reports have demonstrated that innate immune response and inflammation have an important role in major neurodegenerative diseases. The activation of the NF-κB family of transcription factors is a key step in the regulation of pro inflammatory cytokine expression. Microglia and other cell types in the brain can be activated in response to endogenous danger molecules as well as aggregated proteins and brain injury. During the past couple of years several studies reported the role of cystatins in neuroinflammation and neurodegeneration. In the present review, I will summarize and analyze recent findings regarding the role of cystatins in inflammation and NF-κB activation. Type I cystatin stefin B (cystatin B) is an endogenous cysteine cathepsin inhibitor localized in the cytosol, mitochondria and nucleus. Mutations in the gene of stefin B are associated with the neurodegenerative disease known as Unverricht-Lundborg disease and microglial activation plays an important role in the pathogenesis of the disease. Stefin B deficient mice have increased caspase-11 expression and secreted higher amounts of pro-inflammatory cytokines. The increased caspase-11 gene expression, was a consequence of increased NF-κB activation.
    Full-text · Article · Dec 2015
    • "The mammalian cystatins belonging to this type are the kininogens (Ohkubo et al., 1984). Cystatins in immune cells have been reported to participate in the release of nitric oxide, phagocytosis, and expression of cytokines (Kopitar-Jerala, 2006; Magister and Kos, 2013; Maher et al., 2014a). "
    [Show abstract] [Hide abstract] ABSTRACT: Stefin B (cystatin B) is an endogenous cysteine cathepsin inhibitor localized in the cytosol, mitochondria and nucleus. Its expression is upregulated upon macrophage activation and cellular stress. Mutations in the gene of stefin B are associated with the neurodegenerative disease known as Unverricht-Lundborg disease (EPM1). It was reported that early microglial activation precedes neuronal loss in the brain of the stefin B-deficient mice, implying a role of the inhibitor at the cross-talk between microglia and cerebellar cells. Detailed analysis of microglial activation in stefin B-deficient microglia showed a significantly higher proportion of both pro-inflammatory M1 and anti-inflammatory M2 microglia in stefin B-deficient mouse brain compared with control mice. In our recent work, we demonstrated that stefin B-deficient mice were significantly more sensitive to the lethal lipopolysaccharide (LPS)-induced sepsis, due to increased caspase-11 expression and secreted higher amounts of pro-inflammatory cytokines IL-1β and IL-18. Upon LPS stimulation, stefin B was targeted into the mitochondria, and the lack of stefin B resulted in the increased destabilization of the mitochondrial membrane potential and mitochondrial superoxide generation. The increased caspase-11 gene expression and better pro- inflammatory caspase-1 and -11 activation determined in stefin B deficient bone marrow-derived macrophages resulted in enhanced non-canonical inflammasome activation. Since signaling pathways in macrophages could be compared to the ones in microglia we propose that inflammasome activation could play an important role in the pathogenesis of EPM1.
    Full-text · Article · Dec 2015
Show more