Download PrPc expression, Dusseldorf, Octobre 2005 - ORBi

Characterization of bovine and human cellular prion protein expressed in the central
nervous system and in lymphoid organs
V. Defaweux1, S. Capellari2, S. Stramiello2, N. Antoine3, G. Dorban1, C. Demonceau1, O. Jolois1, E. Heinen1 and P. Parchi2.
1Dpt
of Morphology and Immunology, Institute of Human Histology, Faculty of Medecine, University of Liège, Belgium–www.ulg.ac.be/histohum.
2Department of Neurological Sciences, Faculty of Medicine, University of Bologna, Italy.
of Animal Histology, Department of Morphology and Pathology, Faculty of Veterinary Medecine, University of Liège, Belgium.
3Laboratoy
Prion cell tropism significantly varies among animal species, depending on both the agent strain and host-specific factors. For
example, prions show high lymphotropism in scrapie infected sheep and in vCJD, but little, if any, in sCJD or BSE. In particular, the BSE
strain is associated with significant PrP-res accumulation in tonsils, spleen and appendix in humans, whereas, it is largely confined to the
nervous system in infected cattle. So, it appears that, at least in the case of BSE and vCJD, host properties can influence the accumulation of
the infectious agent in lymphoid organs.
Given that the normal cellular prion protein (PrPc), is sine qua non for PrP-res formation and the development of TSE, it appears
reasonable to hypothesize that tissue-specific PrPc properties may represent one of the host factors influencing the cell tropism of the
infectious agent in human or bovine.
Western blot analysis of truncated
PrPc forms expressed in bovine CNS
Western blot analysis to compare the ratio of PrPc glycoforms expressed
in the CNS of bovine and human
23
80
SAF32
 α
100
100
 α
231
PrPc
Diglycos.
Monoglycos.
Unglycos.
Bovine medulla
66,62 %
16,76 %
16,62 %
Bovine
cerebellum
73,79 %
15,45 %
10,76 %
80
PrPc
Diglycos.
Monoglycos.
Unglycos.
Human
medulla
59, 02 %
22,38 %
18,59 %
Human
cerebellum
62,08 %
26,32 %
11,60 %
SAF60
23
231
FL (Full length, N-C) (27kDa)
(α-C) (19kDa)
FL (Full length, N-C) (27kDa)
60
(α-C) (19kDa)
100
PrPc
(-C) (21kDa)
80
%
%
(-C) (21kDa)
60
40
Full length
20 kDa frag.
18 kDa frag.
Bovine medulla
66,10 %
11,81 %
28,10 %
Bovine
cerebellum
78,42 %
6,31 %
15,28 %
40
60
20
0
0
32KDa
%
20
40
Diglycosylated
PrPc
Monoglycosylated
PrPc
-
Bovine cerebellum
+
Diglycosylated
PrPc
n=8
Bovine medulla
PNGaseF
Unglycosylated
PrPc
n=4
P value <5%
20
0
P value <5%
Full Length
18KDa
20 KDa
fragment
18 KDa
fragment
100
PNGaseF
80
80
60
60
40
40
20
20
0
0
Diglycosylated
PrPc
Monoglycosylated
PrPc
n=7
n=8
Human medulla
Bovine medulla
Diglycosylated
PrPc
Unglycosylated
PrPc
P value <5%
Monoglycosylated
PrPc
Human Cerebellum
n=7
Bovine Cerebellum
n=4
+
Med S6381
+
Cer S6381
Bovine
+ medulla - n = 8 +
Bovine cerebellum
n=4
Med S6380
Cer S6380
P value <10%
P value <5%
 The expression of truncated forms of
PrPc (i.e. 21 and 18 kDa PrPc) is also
significantly heterogenous according to the
brain region investigated.
%
%
 PrPc
glycoform
ratios
are
significantly
different
between
cerebellum and medulla in bovine and
human.
 Only the unglycosylated PrPc is
distributed like wise in medulla and in
the cerebellum of bovine and human.
26KDa
Unglycosylated
PrPc
n=7
n=7
Human medulla
Human cerebellum
100
Cer s6375
Monoglycosylated
PrPc
Unglycosylated
PrPc
P value <5%
Western blot analysis to compare the PrPc glycoform ratios and the truncated PrPc forms expressed
in bovine lymphoid tissues
in bovine lymhpoid cells
 α
 α
23
SAF32
SAF60
231
23
FL (Full length, N-C) (27kDa)
Isolation
of
PrPc
expressing
follicular
dendritic cells (FDC)
231
(α-C) (19kDa)
FL (Full length, N-C) (27kDa)
(-C) (21kDa)
(α-C) (19kDa)
(-C) (21kDa)
 α
Medullar
PNGase F
-
+
Spleen
Fo
To
+
+
-
Fo
Mln
23
SAF32
231
FL (Full length, N-C) (27kDa)
(α-C) (19kDa)
+
FDC ultrastructure SAF32+ FDC
(-C) (21kDa)
32KDa
Jejunal Peyer’s Patches
Tonsils
Mesenteric lymph nodes
32KDa
32KDa
26KDa
32KDa
26KDa
32KDa
26KDa
26KDa
PNGaseF
18KDa
26KDa
PNGaseF
PNGaseF
Medul
+
Medul
+
Spleen
+
Fo
MLN
-
+
Total Spleen
+
Fo
To
+
Lymphocytes
-
+
Lymphocytes
-
+
Follicular dendritic cells
Fo
-
+
Lymphocytes
-
+
Follicular
Dendritic cells
+
Lymphocytes
-
+
Follicular
Dendritic cells
 PrPc is highly glycosylated in spleen and in lymphoid follicles isolated from bovine lymphoid tissues as well as in their FDC and lymphocytes.
 After deglycosylation, a novel PrPc truncated form with a relative molecular mass of about 25 kDa was detected in bovine lymphoid organs
beside the typical 18 and 21 kDa forms.
Western blot analysis to compare the PrPc glycoform ratios and the
truncated PrPc forms expressed in human lymphoid tissues
Human Tonsil
Human spleen
 α
32KDa
23
SAF32
231
FL (Full length, N-C) (27kDa)
(α-C) (19kDa)
26KDa
(-C) (21kDa)
-
+
PNGaseF
-
+
Follicles
-
+
Lymphocytes
-
+
Follicular
Dendritic
Cells
 No difference in WB PrPc profile was seen in follicles, lymphocytes and FDC of
human tissues
 Immune PrPc is highly glycosylated and, after deglycosylation, the 25 kDa is also
expressed in human lymphoid tissues and cells.
Our results highlight variation in
the profile expression of PrPc in
peripheral and central tissues of
bovine
and
human.
Such
differences
may
have
an
implication for PrPc function and
may represent critical factors
influencing the accumulation of the
infectious agent in these areas.
Supported by the EU contract QLG3-CT-2002-81030