Low activity of complement in the cerebrospinal fluid of the patients with various prion diseases
- Cao Chen†1, 2,
- Yan Lv†1, 2,
- Qi Shi1, 2,
- Wei Zhou1, 2,
- Kang Xiao1, 2,
- Jing Sun1, 2,
- Xiao-Dong Yang1, 2 and
- Xiao-Ping Dong1, 2, 3Email author
© Chen et al. 2016
Received: 12 August 2015
Accepted: 7 April 2016
Published: 3 May 2016
The aim of this study was to analyze the state of activity and levels of complement in the cerebrospinal fluid (CSF) of patients with various prion diseases (PrDs).
The proteomic data emphasized the levels of 20 known complement components found in the CSF of the sCJD panel that were lower than those found in the non-PrD panel. 50 % of the complement hemolytic activity (CH50) assays revealed significantly lower activity of complement in the CSF of the sCJD panel. The decreased levels of three key complement subunits, C3a/α, C4β, and C9 in the CSF of the sCJD panel were verified by Western blots. Furthermore, the CH50 values in the CSF of 136 sCJD, 39 gCJD, 22 FFI and 145 non-CJD patients were individually tested. Compared with the control of non-PrD, the CH50 value in the CSF specimens of various PrDs, especially in three subtypes of inherited PrDs, were significantly lower. Relationship analysis identified that the CH50 activity in the CSF was negatively associated with the protein 14–3–3 positive in the CSF.
These results indicate a silent complement system in the CSF of PrD patients.
Please see Additional file 1 for translations of the abstract into the six official working languages of the United Nations.
Human prion diseases (PrDs) are a category of fatal neurodegenerative disorders including Creutzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Scheinker syndrome (GSS), fatal familial insomnia (FFI) and Kuru . Surrogate biomarkers in CSF are screened for alterations, which not only identify the diagnostic markers for CJD, but also provide insight for understanding the pathogenesis. So far, only the immunoblot for the 14–3–3 protein in CSF is included in the diagnostic criteria for sporadic CJD (sCJD) .
The complement system, which consists of at least 30 kinds of soluble and membrane-bound proteins, is an immune-defense mechanism with a wide range of effects that take place in the central nervous system (CNS). Evidence suggests that the activation of complement components occurs in patients with various neurodegenerative disorders [3–6], and that certain complement proteins are deregulated during prion infection [5, 7–12]. Our previous proteomic study has revealed that some complement components were lower in the CSF of sCJD patients . However, the exact relationship has yet to be proven.
This study was approved by the Ethical Committee of the National Institute for Viral Disease Control and Prevention, China CDC, under protocol 2009ZX10004–101.
The relevant characteristics of the patients with non-CJD and sCJD, gCJD and FFI
Median age at onset (range) (y)
47.5 (19–63) a
Age at onset <50 years (%)
Age at onset 50–70 years (%)
Age at onset >70 years (%)
Patients with PRNP gene sequenced (%)
Codon 129 genotype
EEG Typical/Total (%)
92/136 (67.6) b
8/14 (57.1) b
14–3–3 Positive/Total (%)
99/136 (72.8) b
18/25 (72) b
14/14 (100) b
Progressive dementia/Total (%)
128/136 (94.1) b
106/136 (77.9) b
15/25 (60) b
10/14 (71.4) b
12/22 (54.5) b
Visual or cerebellar disturbance (%)
88/136 (64.7) b
18/25 (72) b
11/14 (78.6) b
11/22 (50) b
Pyramidal or extramidal dysfunction (%)
104/136 (76.5) b
12/14 (85.7) c
Akinetic mutism (%)
66/136 (48.5) b
8/14 (57.1) b
All enrolled CSF samples were obtained by standard clinical procedures and were free of blood contamination. Routine CSF biochemistry assays of those specimens, including cell count, glucose, and total protein were all within the normal ranges.
The detection of the 50 % of complement hemolytic activity (CH50, U/ml) in CSF (50 μl of each tested sample) was derived from the commercial double-antibody sandwich enzyme-linked immunosorbent assays (ELISAs, YAD, China). Western blots for the complement components C3a/α, C4β and C9 in the panels of the pooled CSF (20 μl of each tested sample) were conducted using anti-C3a/α mAbs (sc-47688), anti-C4β pAbs (sc-25816) and anti-C9 mAbs (sc-390000) (Santa Cruz biotechnology Inc). The detailed procedures of CH50 ELISA and Western blot are described elsewhere . The data was processed with GraphPad Prism 5 and SAS 9.2 statistics software.
The numbers of CSF samples among different human prion diseases according to various CH50 value ranges
CH50 (U/ml) Median (min, max)
CH50 (U/ml) % (case No./group No.)
28.3 % (41/145)
29.7 % (43/145)
24.1 % (35/145)
17.9 % (26/145)
49.3 % (67/136)
38.2 % (52/136)
7.4 % (10/136)
5.1 % (7/136)
92 % (23/25)
8 % (2/25)
100 % (14/14)
77.3 % (17/22)
22.7 % (5/22)
Multivarivate logistic regression of the levels of CSF CH50 and relational influence factors
OR 95 % (CI)
0.793 (0.456, 1.376)
Age of onset
1.260 (0.343, 4.623)
1.503 (0.713, 3.169)
Codon 129 genotype
1.114 (0.107, 11.648)
0.231 (0.127, 1.003)
0.684 (0.348, 1.346)
0.372 (0.195, 0.707)
1.104 (0.485, 2.510)
1.137 (0.588, 2.199)
Visual or cerebellar disturbance
0.639 (0.340, 1.202)
Pyramidal or extramidal dysfunction
0.602 (0.335, 1.082)
0.692 (0.337, 1.419)
multivarivate logistic regression
0.362 (0.182, 0.722)
This study has been the first time that lower CSF complements in PrD patients have been observed. The changes in complement components in CSF have either decreased  or increased . However, the CSF samples in those studies are not from a healthy control group, but from patients with various neurological disorders. Activation of the complement system is frequently detected in the brains of PrD patients. The reason for this remains unclear, in addition to the relatively silent complement. It might be assumed that the recruitment of the large amount of complement components in the brain [3, 5], such as forming C1q-PrPSc complex [16, 17] and membrane attack complex (MAC) , might lead to smaller amounts of complement penetrating CSF.
Unlike the wide-range distribution of CSF CH50 values in sCJD, the CH50 values in the three subtypes of genetic PrDs are low with a narrow range. It indicates a silent state of the innate immune system in the central nervous system during the pathogenesis of genetic PrDs. In accordance with our previous study, microglia and some cytokines are silent in the post-mortem brains of FFI and G114V gCJD cases, while some are obviously activated in other types of sCJD cases . Recent iTRAQ-based proteomic study of the brain tissues of sCJD, FFI, and G114V gCJD show multiple pathways involving immunity and infection in the brains of sCJD subjects, except for hereditary cases . Additionally, the diversity of clinical and pathological characteristics in sCJD may also be contributing factors to the variation in CH50 present within CSF.
14–3–3 positive CSF samples are the unique factor correlated with CH50 values found in the CSF of sCJD patients. The exact molecular association between these two factors is unknown. Contrary to the well-known diagnostic evidence for sCJD, the correlation of 14–3–3 CSF samples with brain pathology in CJD are rarely reported, and draw controversial conclusions [20–22]. 14–3–3 samples are also associated with the alterations of other proteins in the CSF of sCJD patients. Our recent publication illustrates that higher levels of tau isoforms containing Exon-2 and Exon-10 segments in sCJD patients with 14–3–3 positive CSF . The NSE level in CSF of sCJD patients is also associated with 14–3–3 positive readings . In that case, one might assume that 14–3–3 positive CSF samples of sCJD patients correlate with the alterations of CH50 values and other biomarkers in CSF, thus representing a (sCJD) disease-associated feature. Nevertheless, the exact molecular mechanism of 14–3–3 positive and lower CH50 levels in the CSF of sCJD patients deserves further study.
The complement activities in the CSF of patients with prion diseases (including sCJD, gCJD, and FFI), are lower than their non-CJD counterparts are. Additionally, subjects whose CSF samples tested 14–3–3 positive exhibit lower complement activity in sCJD patients.
This work was supported by Chinese National Natural Science Foundation Grants (81401670), China Mega-Project for Infectious Disease (2011ZX10004-101, 2012ZX10004215) and the SKLID Development Grant (2012SKLID102).
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