SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE

ALZHEIMER’S DISEASE Copyright © 2020

The Authors, some
Chi3l1/YKL-40 is controlled by the astrocyte circadian rights reserved;
exclusive licensee
clock and regulates neuroinflammation and Alzheimer’s American Association
for the Advancement
disease pathogenesis of Science. No claim
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Brian V. Lananna1, Celia A. McKee1, Melvin W. King1, Jorge L. Del-Aguila2, Julie M. Dimitry1,
Fabiana H. G. Farias2, Collin J. Nadarajah1, David D. Xiong1, Chun Guo3, Alexander J. Cammack1,
Jack A. Elias4, Jinsong Zhang3, Carlos Cruchaga2,5, Erik S. Musiek1,5*

Regulation of glial activation and neuroinflammation are critical factors in the pathogenesis of Alzheimer’s disease
(AD). YKL-40, a primarily astrocytic protein encoded by the gene Chi3l1, is a widely studied cerebrospinal fluid
biomarker that increases with aging and early in AD. However, the function of Chi3l1/YKL-40 in AD is unknown. In

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a cohort of patients with AD, we observed that a variant in the human CHI3L1 gene, which results in decreased CSF
YKL-40 expression, was associated with slower AD progression. At baseline, Chi3l1 deletion in mice had no effect
on astrocyte activation while modestly promoting microglial activation. In a mouse APP/PS1 model of AD, Chi3l1
deletion decreased amyloid plaque burden and increased periplaque expression of the microglial lysosomal marker
CD68, suggesting that Chi3l1 may suppress glial phagocytic activation and promote amyloid accumulation.
Accordingly, Chi3l1 knockdown increased phagocytosis of zymosan particles and of -amyloid peptide in both
astrocytes and microglia in vitro. We further observed that expression of Chi3l1 is regulated by the circadian clock,
as deletion of the core clock proteins BMAL1 or CLOCK/NPAS2 strongly suppresses basal Chi3l1 expression, whereas
deletion of the negative clock regulators PER1/PER2 increased Chi3l1 expression. Basal Chi3l1 mRNA was non-
rhythmic because of a long mRNA half-life in astrocytes. However, inflammatory induction of Chi3l1 was gated by
the clock. Our findings reveal Chi3l1/YKL-40 as a modulator of glial phagocytic activation and AD pathogenesis in
both mice and humans and suggest that the astrocyte circadian clock regulates inflammatory Chi3l1 induction.

INTRODUCTION from oxidative stress in the periphery (12–14). In AD, numerous

Neuroinflammation plays a critical role in the pathogenesis of most
neurodegenerative diseases, including Alzheimer’s disease (AD) (1, 2).
In AD, triggering of the brain’s innate immune response, character-
ized, in part, by activation of astrocytes and microglia, can exert both
degenerative and protective effects in a context-dependent manner
(1, 3). Although astrocytes and microglia surround amyloid plaques
and limit plaque growth through -amyloid (A) phagocytosis, deg-
radation, and clearance, they can also exacerbate pathology via
dysfunctional inflammatory responses (1, 2). Thus, a deeper under-
standing of factors affecting glial function and neuroinflammation
in neurodegenerative conditions is needed.
YKL-40, a secreted glycoprotein encoded by the Chi3l1 gene, is a
well-described human cerebrospinal fluid (CSF) biomarker of neuro­
inflammation, which is elevated in AD (4–6), as well as other
neurologic diseases including multiple sclerosis, amyotrophic lateral
sclerosis, and frontotemporal dementia (7–9). CSF YKL-40 expres-
sion steadily increases with age starting in middle age, even in
amyloid-negative individuals (5). Chi3l1/YKL-40 is expressed pri-
marily in astrocytes in the brain, in macrophages in the periphery,
and is induced in the setting of inflammation (4, 10, 11). Animal
studies suggest that it may mitigate inflammation and protect cells
1
Department of Neurology, Washington University School of Medicine, St. Louis,
MO 63110, USA. 2Department of Psychiatry, Washington University School of Med-
icine, St. Louis, MO 63110, USA. 3Department of Pharmacological and Physiological
Science, Saint Louis University School of Medicine, St. Louis, MO 63104, USA. 4Division
of Medicine and Biological Sciences, Brown University, Providence, RI 02903, USA.
5
Knight Alzheimer’s Disease Research Center and Hope Center for Neurological
Disease, Washington University School of Medicine, St. Louis, MO 63108, USA.
*Corresponding author. Email: musieke@wustl.edu
studies show that CSF Chi3l1/YKL-40 increases in parallel with tau
and other markers of inflammation and neurodegeneration and that
elevated Chi3l1/YKL-40 predicts disease progression (4, 5). Although
Chi3l1/YKL-40 has been studied extensively as a biomarker, little is
known about its function in the brain and in AD.
Many patients with neurodegenerative diseases also exhibit
circadian system dysfunction, including those with preclinical AD
(15). This dysfunction has been implicated as a potential contributor
to AD pathogenesis (16, 17). The circadian system orchestrates
24-hour rhythms in many physiological and behavioral parameters,
including sleep, activity, and endocrine function (18). The core cir-
cadian clock consists of a transcription-translation feedback loop,
driven by the master circadian transcription factor BMAL1 (Brain and
Muscle ARNT-Like 1), which regulates circadian rhythms in tran-
scription in a cell type–specific manner. The circadian clock regulates
immune responses in a variety of peripheral innate and adaptive im-
mune cell types (19). Astrocytes and microglia also have functioning
circadian clocks, and circadian timing can affect their inflammatory
responses (20–22). Our group has shown that disruption of core
circadian clock transcription via deletion of Bmal1 can promote
neuroinflammation and that BMAL1 regulates astrocyte activation
in a cell-autonomous manner (23, 24). However, the mechanisms
linking glial clocks to neuroinflammation are poorly understood.
Here, we report data from humans and mouse models suggest-
ing that Chi3l1/YKL-40 accelerates AD pathogenesis, potentially by
altering glial function in the brain. We also show evidence from mice
that the core circadian clock in astrocytes strongly regulates Chi3l1
transcription and gates its inflammatory induction. Together, our
results reveal Chi3l1/YKL-40 as a modulator of AD pathogenesis

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SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE

and illuminate a link between glial circadian clocks and Chi3l1
expression.
RESULTS
A human genetic variant causing decreased CHI3L1/YKL-40
is associated with slower disease progression in human
patients with AD
We sought to determine whether CHI3L1/YKL-40 influences AD
pathogenesis in humans. First, we examined a human single-nucleus
RNA sequencing (RNAseq) dataset derived from cortical brain
tissue from three patients with AD (25) and observed that CHI3L1
expression was strongly associated with the astrocyte cell cluster
(Fig. 1A). Although a few CHI3L1-expressing microglia were present,
differential gene expression analysis shows that CHI3L1 is enriched
ated with any changes in AD progression. We examined data from
778 participants enrolled in longitudinal observational studies and
included only participants with AD (confirmed by clinical assess-
ment and CSF biomarker profile), 26% of whom carried the CC_TT
polymorphism at rs10399931. We examined clinical progression of
AD as assessed by the rate of increase in the Clinical Dementia Rating
Sum-of-Boxes score (CDR-SB). We found that the rs10399931 SNP
was significantly associated (P = 0.031) with a slower rate of AD
progression, suggesting that people with genetically lower CHI3L1/
YKL-40 expression have 16% slower disease progression (Fig. 1C).
For comparison, pathogenic variants in the Trem2 gene were previ-
ously associated with a 23% increased rate of progression using the
same methods (27). This initial human data suggested that CHI3L1/
YKL-40 might be a modulator of human disease pathogenesis. Thus,
we sought to examine the mechanisms by which CHI3L1/YKL-40

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in astrocytes and minimally expressed in neurons (Fig. 1B). We
have previously reported that CSF YKL-40 expression in humans is
strongly controlled by genetic variation in the CHI3L1 locus and
have described a common genetic variant in the CHI3L1 locus
(rs10399931) associated with decreased CSF YKL-40 concentrations
(26). Using the Knight Alzheimer’s Disease Research Center at
Washington University clinical database, we next examined whether
this rs10399931 single-nucleotide polymorphism (SNP) was associ-
might exacerbate AD pathogenesis, including glial activation, in
mouse models.
Chi3l1 modulates the astrocytic and microglial inflammatory
responses in vivo
Our data (Fig. 1A) and other studies show that Chi3l1 is highly en-
riched in human astrocytes (28) and only negligibly expressed in
other cell types, whereas in the mouse central nervous system, at

A 40
B

Differential expression (fold)

5
****
20 4

3
tSNE_2

0 2

1
–20
0
ro e
ur a
O n
PC
N e g li
ic yt

o
M oc
r

–20 0 20 40
st
A

tSNE_1
C
40
CHI3L1 CHI3L1
Rs10399931
CC
15

20
CDR-SB

CC_TT
tSNE_2

10

0 P = 0.031
5

–20

0 2 4 6 8 10
–20 0 20 40 Years
tSNE_1
Fig. 1. A polymorphism in human CHI3L1 affects rate of AD progression. (A) Single-nucleus RNAseq t-distributed stochastic neighbor embedding (tSNE) plot showing
cell clusters (top) or CHI3L1 expression (purple, bottom). Ex, excitatory neuron; In, inhibitory neuron; OPC, oligodendrocyte precursor cells; Endo, endothelial cells. Data
are available at http://ngi.pub/snRNAseq/. (B) CHI3L1 expression by cell cluster from data in (A). ****P = 3 × 10−280. (C) AD progression (change in CDR-SB) from human patients
with and without the CC_TT polymorphism of the rs10399931 SNP in CHI3L1.

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SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE

baseline, it is highly expressed in astrocytes and oligodendrocyte
precursor cells but less so in microglia (29). Thus, we sought to in-
vestigate the role of astrocytic Chi3l1 in regulating neuroinflamma-
tion. We used small interfering RNA (siRNA) to knock down Chi3l1
(siChi3l1) in mouse primary astrocyte cultures, achieving a 91%
decrease in Chi3l1 mRNA (Fig. 2A). Loss of Chi3l1 induced a small
decrease in Gfap and a small increase in Il6. This suggests that
Chi3l1 suppression may exert differential effects on astrocyte acti-
vation and cytokine expression. In vivo, constitutive Chi3l1 knockout
(KO) (30) had no effects on expression of several inflammatory
transcripts (Fig. 2B and fig. S1C). As a group, transcripts related to
astrocyte activation were modestly altered at baseline by Chi3l1 KO,
although no individual transcripts were changed (Fig. 2C and fig. S2A).
Similarly, microglial activation genes (including Trem2, Spp1, Gpnmb1,
and Apoe) were slightly increased at baseline as a group in Chi3l1
hippocampus (Fig. 2, E and F). However, Chi3l1 deletion subtly
increased staining for the microglial marker IBA1 (ionized calcium-­
binding adapter molecule 1) in the cortex (Fig. 2, G and H), suggest-
ing that Chi3l1 differentially modulates astrocyte and microglial
activation at baseline.
We next examined the effect of Chi3l1 on acute neuroinflammation
induced by the inflamogen lipopolysaccharide (LPS). In primary
astrocyte cultures, Chi3l1 siRNA exacerbated LPS-induced cytokine
expression (fig. S1A). Chi3l1 KO mice exhibited a general exacerbation
of the inflammatory response with increased hippocampal expres-
sion of several inflammatory transcripts, including several microglia-­
specific transcripts such as Cybb and Nlrp3 after intraperitoneal LPS
injection (fig. S1, B to E). However, Chi3l1 KO did not alter LPS-­
induced expression of astrocyte activation markers or AD-associated
microglial activation markers (fig. S2, A and B). Together, our data

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KO mice, although no individual genes were different (Fig. 2D and suggest that Chi3l1/YKL-40 deletion did not affect astrocyte activa-
fig. S2B). Chi3l1 did not alter baseline glial fibrillary acidic protein tion but mildly enhances microglial activation at baseline and mod-
(GFAP) immunoreactivity, a marker of astrocyte activation, in the estly potentiates LPS-induced inflammatory cytokine expression in
astrocytes and microglia.

ns
* * Deletion of Chi3l1 reduces amyloid
A B WT KO C WT KO D WT KO plaque deposition in a mouse
Wild-type astrocytes Gfap 3
model of AD

Fold change
Ifng Aif1
mRNA expression (fold)

1.5 * siScr
Lcn2
2
As YKL-40 is increased in the CSF of
Il10
Timp1 Cd68
**** ** siChi3l1 Il12
patients with AD and used as a bio-
Serpina 1
Il1a Gpnmb1
1.0 Serping1
Il1b Gbp2
Psmb8
Spp1 0
marker of AD (4, 6), we next sought to
test the hypothesis that Chi3l1/YKL-40
Il33
0.5 Il6 Amigo2 Tmem119
Tnfa
Ptgs2
S100a6
Trem2 could also influence pathology in a mouse
0.0
Ccl2 Aqp4 Tgfb model of AD-related -amyloidosis. We
crossed amyloid precursor protein (APP)/
Cxcl2 Apoe
Chi3l1 Gfap Il6 Apoe
Cxcl5 Megf10
PS1-21 mice, which express the human
WT Chi3l1 KO
APP gene with the KM670/671NL
E GFAP F (Swedish) mutation and human presenilin 1
Retrosplenial Ctx Hippocampus (PS1) with the L166P mutation (31), with
6 30 P = 0.059
wild-type (WT) or Chi3l1 KO mice. All
GFAP (% area)
GFAP (% area)

mouse brain tissues were harvested at

20
4
8 months of age when mice had devel-
2 10
oped substantial plaque pathology in
both the cortex and hippocampus (31).
0 0
Staining with X34, which selectively
WT Chi KO WT Chi KO labels -pleated sheet fibrillar A plaques
G IBA1 H (32), revealed that loss of Chi3l1 reduced
fibrillar plaque number by 21% (Fig. 3,
Retrosplenial Ctx Hippocampus
A and C) and plaque area by 17% (Fig. 3,
25 * 16 A and D) in the hippocampus but did
not alter these measures in the cortex
% IBA1 area

% IBA1 area

20
12
15
8
(fig. S3, A to C). Staining with an A anti-
10 body (HJ3.4) revealed a much more pro-
5
4
nounced 55% reduction in plaque burden
0 0 in the hippocampus (Fig. 3, A and E)
WT Chi KO WT Chi KO
and a 42% decrease in the cortex (fig. S3,
Fig. 2. Loss of Chi3l1 mildly shifts glial activation. (A) qPCR gene expression from primary astrocytes transfected A and D). This discrepancy between
with control (siScr) or Chi3l1 (siChi3l1) siRNA. n = 6 to 10 replicates from three independent experiments. (B to D) Cytokine
staining methodologies led us to hypoth-
and chemokine (B), astrocyte activation marker (C), or microglia activation marker (D) expression from Fluidigm qPCR
esize that Chi3l1 deletion results in the
of 2- to 5-month-old Chi3l1−/− and WT control mouse cortex 6 hours after intraperitoneal PBS. Mean of six mice per
group normalized to WT. Two-way ANOVA with Tukey correction for multiple comparisons. ns, not significant. (E to
selective reduction of nonfibrillar A.
H) Representative images depicting GFAP (astrocyte) staining (E) and associated quantification (F) or IBA1 (microglia) Subtraction of X34 signal from total plaque
staining (G) and associated quantification (H) in Chi3l1−/− and WT control mice. Scale bars, 400 m. All data represent (HJ3.4) staining indeed revealed a halo
means ± SEM. *P < 0.05, **P < 0.01, and ****P < 0.0001 by two-tailed Student’s t test with Holm-Sidak correction for of aggregated, nonfibrillar A surround-
multiple comparisons when appropriate. ing the fibrillar plaque core in a majority

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SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE

A X34 HJ3.4 Merge HJ3.4-X34

WT:APP/PS1
Chi3l1 KO:APP/PS1

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B C D G H
WT:APP/PS1 ChiKO:APP/PS1 WT:APP/PS1 ChiKO:APP/PS1
X34 plaque number X34 plaque area
150 2.0 *
X34+ plaque number

*
% X34 area

1.5
100
1.0

X34 50
0.5

0 0.0
WT Chi KO WT Chi KO

E F
HJ3.4 plaque area Nonfibrillar plaque
HJ3.4 8 ***
*** 5
HJ3.4-X34 area
% HJ3.4 area

6
4

×
3
4
2
2
1

0 0
HJ3.4-X34
WT Chi KO WT Chi KO
Fig. 3. Loss of Chi3l1 mitigates amyloid pathology. (A) Representative hippocampal images from 8-month-old Chi3l1−/−:APP/PS1+ and APP/PS1+ control mice depict-
ing staining by X34 (fibrillar plaques), HJ3.4 antibody (total A), and subtraction of fibrillar (X34) from total (HJ3.4) A. The hippocampus is outlined in yellow. The yellow
rectangle denotes region of inset in (B). Scale bar, 300 m. (B) Representative higher magnification images depicting stains from (A). “Halo” of nonfibrillar A surrounding
fibrillar plaque core (X34) substantially reduced in Chi3l1−/− mice. Scale bar, 50 m. (C to F) Quantification of X34+ puncta (fibrillar plaque number) (C), X34+ area (fibrillar
A) (D), HJ3.4+ area (total A) (E), or area covered by nonfibrillar A (total A in HJ3.4-fibrillar X34) (F) in hippocampal staining from (A). (G) X34 and HJ3.4 colocalization
(top) with 3D surface rendering of X34 (red) and HJ3.4 (green) staining (middle) with 20-m shells (pink) around each fibrillar plaque. Scale bar, 30 m. (H) Quantification
of nonfibrillar plaque volume as ratio of HJ3.4 to X34 (top) or total A in HJ3.4-fibrillar X34 (bottom) in 20-m shell surrounding each X34+ plaque in a subset of mice from
(C) to (F). Data points represent average of two to four sections per mouse and 4 to 6 (Chi3l1+/+) and 4 to 12 (Chi3l1−/−:) mice per group. All data represent means ± SEM.
*P < 0.05, **P < 0.01, and ***P < 0.001 by two-tailed Student’s t test.

of plaques in control APP/PS1 mice, as well as some X34-negative,
HJ3.4+ plaques (Fig. 3, A and B, and fig. S3A). We observed an 85%
loss of this nonfibrillar A in the hippocampus of Chi3l1 KO mice
(Fig. 3F), along with a 54% reduction in the cortex (fig. S3E). To more
closely investigate this selective loss of nonfibrillar A, regions with
similar amounts of fibrillar (X34+) plaque load (fig. S3, F and G) were
imaged using confocal microscopy and three-dimensional (3D) re-
construction in a subset of mice. Plaque volume measurements again
revealed a loss in antibody (HJ3.4)–positive A (fig. S3H) and con-
firmed the selective loss of nonfibrillar A in the absence of Chi3l1
(Fig. 3, G and H). Moreover, loss of Chi3l1 altered the distribution of
antibody (HJ3.4)–stained plaques in APP/PS1 mice (fig. S4, A and B)
with a substantial decrease in plaque number and a small decrease
in average plaque area (fig. S4, C and D). This pool of aggregated but
nonfibrillar A could not be separated biochemically via sequential
fractionation with increasing concentrations of guanidine (fig. S4E).
Loss of Chi3l1 did not result in changes in the amount of APP protein
(fig. S4F) or in the expression of several key amyloid processing/
metabolic genes, including App, Bace1, Ide, Mmp2 or Mmp9, Ldlr,
Lrp1, Mme, Klk7, and Apoe (fig. S4G). These data, in combination

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with the fact that Chi3l1 is generally not expressed in neurons, indicate
that YKL-40 likely does not alter the production of A. Together,
these data suggest that the loss of Chi3l1 mitigates the accumulation
of A plaque pathology, particularly nonfibrillar plaque material.
Periplaque astrocyte activation is suppressed by
Chi3l1 deletion
As astrocytes and microglia have been shown to play integral roles
in both the clearance of and neuroinflammation induced by A (1, 2),
we next examined the consequences of Chi3l1 deletion on the glial
response to A plaques. Whereas previous reports disagree about
whether impairing astrocyte activation increases or decreases A
plaque burden (33, 34), we found that Chi3l1 deletion resulted in
decreased astrocyte activation as measured by GFAP staining in the
hippocampus and throughout the cortex in APP/PS1+ mice (Fig. 4,
A and B, and fig. S5A). Modest periplaque GFAP reductions were
also observed (Fig. 4, A and C), as controlling for the decreased
fibrillar plaque area (Fig. 4C) or number (fig. S5B) could not fully
account for the lessened GFAP, particularly in the hippocampus.
Thus, the amount of GFAP staining per plaque area is reduced with
Chi3l1 deletion. In concordance with our staining data, transcriptional
profiling of astrocyte activation markers revealed a slight attenuation

A B 50 *** ** *

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40
WT
WT:APP/PS1

Chi KO

% GFAP area
30
X34 GFAP Merge
20

10

0
Hippocampus RS cortex Motor cortex

C
Chi3L1 KO:APP/PS1

35 * P = 0.074
% GFAP area/X34 plaque area

30

25
WT
Chi KO
20

15

10

0
Hippocampus RS cortex Motor cortex

D E F
Fold change

Fig. 4. Loss of Chi3l1 mitigates astrogliosis but facilitates phagocytosis in the presence of A. (A) Representative high-magnification images from hippocampi of
8-month-old Chi3l1−/−:APP/PS1+ and APP/PS1+ control mice stained for X34 (fibrillar plaques) and GFAP (astrocytes). Scale bars, 20 m. (B and C) Quantification of GFAP
coverage (B) or GFAP coverage normalized to X34+ area in the same section (C) from mice in (A). Quantified from wide-field image in fig. S5. RS, retrosplenial. n = 6
(Chi3l1+/+) and 12 (Chi3l1−/−:) mice per group. (D) Astrocyte activation marker gene expression from Fluidigm qPCR of 8-month-old Chi3l1−/−:APP/PS1−, WT:APP/PS1−,
Chi3l1−/−:APP/PS1+, and APP/PS1+ control mouse hippocampus. Mean of 4 mice (APP/PS1−) or 6 to 10 (APP/PS1+) mice per group normalized to WT:APP/PS1−. Two-way
ANOVA with Tukey correction for multiple comparisons. (E and F) pHrodo-labeled zymosan bead (E) or TAMRA-A (F) uptake by primary astrocyte cultures transfected
with control (siScr) or Chi3l1 (siChi3l1) siRNA, ±cytochalasin D to inhibit phagocytosis (+cytoD). Each point represents one field of view with an average of 804 (E) or 517
(F) cells per field. Data are from two independent experiments. All data represent means ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, analyzed by two-
tailed Student’s t test (B and C) or one-way ANOVA (E and F). All data were subjected to Sidak correction for multiple comparisons unless otherwise noted.

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SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE

of hippocampal astrocyte activation in Chi3l1 KO;APP/PS1 mice
(Fig. 4D and fig. S5C). Chi3l1 mRNA was increased in APP/PS1
cortex at 8 months and was absent in Chi3l1 KO mice (fig. S5C).
As astrocyte activation state can affect phagocytosis (3) and
astrocytes are known to phagocytose A plaque material (35), we next
sought to evaluate whether Chi3l1 may be regulating phagocytosis
in astrocytes. siRNA-mediated Chi3l1 knockdown, which suppresses
Chi3l1 mRNA expression in primary astrocyte cultures by ~91%
(see Fig. 2A), increased the phagocytosis of zymosan-coated pHrodo-­
labeled beads by 13% (Fig. 4E) and fluorescent [TAMRA (carboxy-
tetramethyl rhodamine)–labeled] A42 peptide by 50% (Fig. 4F),
each in two separate experiments. In combination, these data support
the idea that loss of Chi3l1 tempers astrocyte activation while poten-
tially increasing astrocytic phagocytosis in response to A plaques.
Chi3l1 deletion promotes plaque-related microglial
CD68 expression
To address the possibility that Chi3l1 could also be regulating the
microglial response to A, we used IBA1 to label microglia and the
microglial lysosomal marker CD68 to assess microglial phagosome
expression (Fig. 5A and fig. S6A). When normalized to plaque area,
we did not observe any changes in absolute coverage of IBA1 in the
hippocampus, motor cortex, or retrosplenial cortex (fig. S6, A and B).
However, the amount of CD68 staining normalized to X34+ plaque
area was elevated across all regions examined in Chi3l1 KO;APP/
PS1 mice (Fig. 5B), indicating increased microglial phagocytic acti-
vation relative to plaque burden. This effect appeared to be driven
by increased CD68 staining around amyloid plaques, as colocalized
IBA1/CD68 area per X34+ plaque area was also increased in Chi3l1

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A B
3.5 ** * *

% CD68 area/X34 area

3.0
WT:APP/PS1

2.5

X34 IBA1 CD68 Merge 2.0

1.5

1.0
WT
Chi KO
0.5

0.0
Hipp RS CTX Motor CTX
C%CD68+ IBA1+ area/X34 area
3.0
Chi3L1 KO:APP/PS1

P = 0.100
* *
2.5

2.0

1.5

1.0
WT
0.5 Chi KO
0.0
ns ** Hipp RS CTX Motor CTX
APP/PS1– APP/PS1+
D WT KO WT KO
E F
Aif1
Cd68
Gpnmb1
Spp1 *
Tmem119
Trem2
6
Apoe
5
Tyrobp 4
Fold change

Lamp1 3
2
Ctsb
1
Tfeb 0

Fig. 5. Loss of Chi3l1 alters microglial activation and enhances A phagocytosis. (A) Representative high-magnification images from hippocampi of 8-month-old
Chi3l1−/−:APP/PS1+ and APP/PS1+ control mice stained for X34 (fibrillar plaques), IBA1 (microglia), and CD68 (phagocytic microglia). Scale bars, 20 m. (B and C) Quantifi-
cation of CD68 (B) or colocalized IBA1/CD68 (C) from mice in (A) normalized to X34+ area in the same section. Quantified from wide-field image in fig. S6. n = 6 (Chi3l1+/+)
and 12 (Chi3l1−/−) mice per group. RS, retrosplenial. (D) Microglia-associated gene expression from Fluidigm qPCR of 8-month-old Chi3l1−/−:APP/PS1−, WT:APP/PS1−,
Chi3l1−/−:APP/PS1+, and APP/PS1+ control mouse hippocampus. Mean of 4 mice (APP/PS1−) or 6 to 10 (APP/PS1+) mice per group normalized to WT:APP/PS1−. Two-way
ANOVA with Tukey correction for multiple comparisons. (E and F) pHrodo-labeled zymosan bead (E) or TAMRA-A (F) uptake by primary microglia cultures transfected with
control (siScr) or Chi3l1 (siChi3l1) siRNA, ±cytochalasin D to inhibit phagocytosis (+cytoD). Each point represents one field of view with an average of 209 (E) or 56 (F) cells
per field. Data are from two (E) or one (F) independent experiments. All data represent means ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 analyzed by
two-tailed Student’s t test (B and C) or one-way ANOVA (E and F). All data were subjected to Sidak correction for multiple comparisons unless otherwise noted.

Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020 6 of 12
SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE
KO mice (Fig. 5, A and C). These changes seem to be CD68 specific
because there were no differences observed in transcript expression
of other lysosomal markers (Fig. 5D and fig. S6C). Transcriptional
analysis of hippocampal tissue for a selection of known microglia
activation markers revealed an increase in the activation marker Spp1
in 8-month-old Chi3l1 KO mice without plaques. We also observed
a slight overall dampening of microglial activation marker expres-
sion (Fig. 5D and fig. S6C) and inflammatory markers (fig. S7, A
and B) with Chi3l1 deletion in the presence of A pathology. These
changes are very likely due to reduced plaque burden in Chi3l1
KO;APP/PS1 mice.
Because of the increase in microglial CD68 expression in Chi3l1
KO;APP/PS1 mice and the observed increase in astrocytic phagocytosis
with loss of Chi3l1, we next examined phagocytosis in cultured pri-
mary mouse microglia. Chi3l1 siRNA (siChi3l1) suppressed Chi3l1
mRNA in microglial cultures by 97% (fig. S7C). Loss of Chi3l1 in
microglia had an even greater effect than in astrocytes, with Chi3l1
knockdown increasing phagocytosis of pHrodo-labeled zymosan
beads by 148% (Fig. 5E) and fluorescent (TAMRA-labeled) A by
100% (Fig. 5F) in microglia in vitro. Together, our data demonstrate
that loss of Chi3l1 leads to decreased amyloid plaque burden and in-
creased microglial CD68 expression in vivo and enhances phagocytosis
of A by both astrocytes and microglia in vitro.
Chi3l1 expression is nonrhythmic but controlled by
the circadian clock
We next explored possible molecular mechanisms regulating Chi3l1
expression and potential link to glial activation. We previously re-
ported that deletion of the master circadian clock gene Bmal1 caused
astrocyte activation in a cell-autonomous manner (24). While ex-
amining an existing transcriptional dataset from control and brain-­
specific Bmal1 KO (Nestin-Cre;Bmal1f/f) cortex to identify circadian
clock targets that may regulate astrocyte activation, we noticed Chi3l1
to be among the most down-regulated transcripts in Bmal1 KO mice.
Although Chi3l1 is reported to be among the 50 most up-regulated
transcripts in activated astrocytes after in vivo inflammation (LPS
injection) (36), our transcriptomic data showed that Chi3l1 was
markedly down-regulated in Bmal1 KO brain (−89%) and strongly
up-regulated in Per1mut;Per2mut (+230%) mice across circadian time
points (Fig. 6, A and B). This unexpected finding of reciprocal ex-
pression changes in mice with mutation of the positive limb (Bmal1)
and negative limb (Per1/2) of the circadian clock suggested that Chi3l1
might be a clock-controlled gene. However, basal Chi3l1 mRNA did
not show circadian oscillation in control mice in this array data
(Fig. 6B). Follow-up quantitative polymerase chain reaction (qPCR)
analysis of cortex tissue collected every 4 hours in constant darkness
confirmed a lack of circadian oscillation in Chi3l1 mRNA in control
mice but again showed an average of a 91% loss in Chi3l1 transcript
in Bmal1 KO brain (fig. S8A). Moreover, we found that, similar to
Bmal1 KO tissue, Chi3l1 mRNA was decreased in cortex from
Clock/Npas2 double-KO mice, as compared to Clock KO alone
(Fig. 6C). Clock/Npas2 double-KO mice lack a binding partner for
Bmal1 and thus have a dysfunctional positive limb of the clock. This
decrease in Chi3l1 expression mirrored that of the BMAL1-CLOCK/
NPAS2 target Nr1d1 (fig. S8B) and occurred despite a compensatory
increase in Bmal1 expression (fig. S8C). These data strongly sup-
port the requirement of a functioning positive limb of the clock
(consisting of BMAL1/CLOCK or BMAL1/NPAS2 heterodimers)
for Chi3l1 expression.
Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020
To test whether the circadian clock is regulating Chi3l1 in astro-
cytes, we measured Chi3l1 in inducible astrocyte-specific Bmal1 KO
(Aldh1l1-CreERT2:Bmal1f/f) mice, which we have previously shown
results in the loss of about 70% of astrocytic BMAL1 (24). Chi3l1
expression was reduced by 52% in the cortex of Cre+ animals (Fig. 6D),
whereas we did not observe any change in Chi3l1 expression in
microglia-specific Bmal1 KO (Cx3cr1-CreERT2:Bmalf/f) mice (Fig. 6E).
Treating primary mouse astrocyte cultures with siRNA targeting
Bmal1 resulted in a 64% loss in the direct BMAL1 transcriptional
target Nr1d1 and a 71% loss in Chi3l1 (Fig. 6F), whereas the secretion
of CHI3L1 protein (YKL-40) decreased by 72% (Fig. 6G). Knockdown
of Bmal1 in primary microglia cultures resulted in a 79% loss in
Nr1d1 but no change in Chi3l1 (fig. S8D). To determine whether
BMAL1 directly regulates Chi3l1, we examined the putative Chi3l1
promoter (within 700 base pairs of the Chi3l1 transcriptional start
Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020
site) and identified six E-boxes, three of which previously displayed
weak binding of BMAL1 in an existing liver chromatin immuno-
precipitation sequencing (ChIP-seq) dataset (fig. S8E) (37). ChIP-qPCR
for BMAL1 binding to three of these E-boxes (fig. S8E) revealed
enrichment when compared to immunoglobulin G (IgG) controls
at all three sites, as well as at a known BMAL1 binding site in the
Dbp promoter (Fig. 6H). Although Chi3l1 transcription is directly
regulated by BMAL1, it is not rhythmic at baseline (Fig. 6B and
fig. S8, A and F). One possibility for the lack of Chi3l1 oscillation in
astrocytes is that its mRNA may have a long half-life. We measured
mRNA degradation kinetics in primary astrocyte cultures after in-
hibition of transcription with actinomycin D and observed that the
half-life of Chi3l1 transcript was much longer than that of Bmal1, a
rhythmic gene (Fig. 6I). Thus, although the positive limb of the cir-
cadian clock directly regulates Chi3l1 expression, basal Chi3l1 mRNA
is not rhythmic likely because of its long half-life.
Induction of Chi3l1 in astrocytes is gated by the 
circadian clock
As Chi3l1/YKL-40 has been shown to increase during inflammatory
conditions (10, 38) and is regulated by nuclear factor B (11), we
next sought to evaluate the regulation of Chi3l1 induction by BMAL1
in the setting of inflammation. In cultured astrocytes, Chi3l1 was
induced by LPS stimulation, but both basal and LPS-stimulated
Chi3l1 expression was suppressed in Bmal1 KO cells (although some
LPS-induced increase in Chi3l1 was still observed) (Fig. 7A). To
more closely examine the possibility that the inflammatory induc-
tion of Chi3l1 is gated by the astrocyte circadian clock, we measured
basal and LPS-induced Chi3l1 expression in synchronized primary
astrocytes across circadian time points. Consistent with our in vivo
data, Chi3l1 transcript did not oscillate in synchronized primary
astrocytes (fig. S8F). However, LPS-mediated induction of Chi3l1
was highly dependent on circadian phase, as LPS-induced Chi3l1
expression varied antiphase to Bmal1 mRNA expression and
closely mirrored the expression pattern of Nr1d1, which is depen-
dent on BMAL1 transcriptional activity (Fig. 7, B and C). This
time-of-day variation in Chi3l1 induction was not observed in cells
treated with Bmal1 siRNA, indicating a requirement for a function-
ing circadian clock (Fig. 7D). These data suggest that circadian
oscillations in BMAL1 transcriptional activity gate the inflam-
matory induction of Chi3l1 in astrocytes (Fig. 7E). The induction of
Chi3l1 in astrocytes was not limited to LPS as exposure of primary
astrocyte cultures to A42 fibrils resulted in an about 4.5-fold in-
crease in Chi3l1 (Fig. 7F).
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A CT6 CT18 B 3 Chi3l1 C Chi3l1 D Astrocytic Bmal1 KO

Chi3l1 mRNA (fold)

1.5

Per1 ,Per2

Per1 ,Per2

Chi3l1 mRNA (fold)

Chi3l1 mRNA (fold)

Bmal1 KO

Bmal1 KO
* 1.25 **
2

m
1.00

Con

Con
1.0
0.75
C4b 0.50
Gfap 1 0.5
A2m 0.25
Serpina3n
Cxcl10 0.00
0.0
Osmr
0 Cre– Cre+
Gbp3
6 12 18 24 KO KO KO KO
S1pr3
s2 ock k al1
ZT a oc
Per1m/Per2m Np Cl /Cl Bm
Steap4
2
Cd44
Endou
Bmal1 f/f as
Np
Slc39a14
Iigp1
Slc22a4
N-Bmal1 KO
B2m

E F G
Aspg
Gsr Microglial Bmal1 KO Wild-type astrocytes Wild-type astrocytes

mRNA expression (fold)

Vim
1.5
*
Nek6 8

YKL-40 (ng/ml)
Chi3l1 mRNA (fold)

Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020

Sulf2
Gbp2 1.25 **** ****
Psmb8 6
1.00 1.0
Tapbp siScr
siBmal1
Sorbs1 0.75
Amigo2 4
Lcn2 0.50 0.5
Olfm1
2
H2-D1 0.25
Igtp
Hspb1 0.00 0.0 0
Cxcl1 Cre– Cre+ Nr1d1 Chi3l1

1
Sc

al
Srgn

Bm
si
Tspan4

si
Cp
H I
Normalized expression

Ifitm3 0.15
H2-T23 IgG control 1.5
Irgm1 Bmal1 Ab ActD

(relative to Actb)
H2-T10

mRNA level
Ggta1
0.10
Gbp10 2.0 1.0
Serping1 Chi3l1
Fold change

Ier3 1.5
Fkbp5
0.05 * Bmal1
Fbln5 0.5
1.0
*
Hspb6
Crispld2
Timp1 0.5 *
Chi3l1 0.00 0.0
0 1 3 8
3
-2
Ch -1

bp
1-
l1
l1

3l

Hours
D
i3
i3

hi
Ch

Fig. 6. Chi3l1 is regulated by the circadian clock. (A) Microarray in Nestin-Cre;Bmal1f/f and Per1/2mut versus control, Bmal1f/f cortex [Lananna et al. (24)] cross-referenced
with 50 genes most up-regulated in astrocytes with in vivo LPS [Zamanian et al. (36)]. CT, clock time. (B) Microarray data from (A) with two additional time points for Cre−
and Per1/2mut mice. (C) qPCR depicting gene expression in global Npas2 KO, Clock KO, Clock/Npas2 double KO, or Bmal1 KO mouse cortex. n = 2 mice per group, normalized
to WT control. (D to F) qPCR showing Chi3l1 expression from Aldh1l1-Cre;Bmal1f/f (ALC) hippocampus (D) or Cx3cr1-Cre;Bmal1f/f (CX3) cortex (E) or gene expression from
WT primary astrocytes 4 to 8 days after transfection with control (siSCR) or Bmal1 (siBmal1) siRNA (F). n = 3 mice per group (D), 6 to 8 mice per group (E), or 15 biological
replicates from five independent experiments (F). (G) Enzyme-linked immunosorbent assay of Chi3l1 protein (YKL-40) in culture medium of WT primary astrocytes 6 days
after transfection with control (siSCR) or Bmal1 (siBmal1) siRNA. n = 3 biological replicates per group. (H) ChIP-qPCR from mouse primary astrocytes with anti-BMAL1
antibody or IgG control. Three distinct E-box–containing regions in the Chi3l1 promoter were assayed. A known BMAL1 binding E-box–containing region in the Dbp
promoter was assayed as positive control. (I) qPCR showing expression of Chi3l1 and Bmal1 mRNA in primary mouse astrocytes treated with actinomycin D at 0 hours and
then harvested at intervals thereafter. n = 3 wells per time point, normalized to Actb mRNA. *P < 0.05 compared to time point 0. All data represent means ± SEM. *P < 0.05,
**P < 0.01, and ****P < 0.0001 by two-tailed Student’s t test with Holm-Sidak correction for multiple comparisons when appropriate.

DISCUSSION Glial activation is thought to be a double-edged sword in AD, as

Because glia can exert either protective or degenerative influences activated glia can phagocytose A and tau and prevent proteopathy,
on the brain, striking a delicate balance of glial activation and in- whereas excessive inflammatory activation can accelerate plaque
flammatory signaling is critical for maintaining brain health. Thus, accumulation and synapse loss (2). Several studies find Chi3l1 to be
factors that alter glial activation state may disrupt this balance anti-inflammatory and/or neuroprotective in the setting of bacterial
and promote neurodegeneration. Although Chi3l1/YKL-40 is an infection (39), traumatic brain injury (14), and experimental
AD biomarker, its role in the progression of AD remains unknown. autoimmune encephalomyelitis (13). However, pharmacologic
Here, we show that in mice and cells, Chi3l1 deletion alters glial inhibition of Chi3l1 has been reported to suppress A deposition in
inflammatory responses, promotes astrocyte and microglial A a rat A infusion model, albeit through a purported anti-inflammatory
phagocytosis, and mitigates amyloid plaque formation. Further- mechanism (40). We observed that Chi3l1 KO enhances inflam-
more, we show that a genetic polymorphism associated with lower mation in response to LPS but not amyloid plaques. Moreover, the
CSF CHI3L1/YKL-40 concentrations in humans is associated with transcriptional signature of microglial activation in response to
slower AD progression. These data suggest that increases in Chi3l1/ LPS versus amyloid plaques is very different, with opposite reg-
YKL-40 that occur during aging and AD (4–6) may have a detri- ulation of key mediators such as Trem2 (1). Thus, the effect of
mental impact on AD pathogenesis by altering glial function and Chi3l1/YKL-40 on the balance of glial activation and neuroinflam-
plaque deposition. mation appears to be context dependent such that loss of Chi3l1

Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020 8 of 12
SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE
Fig. 7. Chi3l1 is induced during inflammation in a Bmal1-dependent manner.
(A) qPCR showing gene expression from WT and Bmal1 KO primary astrocytes ± LPS
(500 ng/ml) for 6 hours. n = 5 independent experiments. (B and C) qPCR showing
LPS-induced Chi3l1 gene expression (B) or expression of Nr1d1 (C) in WT primary
astrocytes synchronized with high serum shock. Cells were treated with LPS (500 ng/ml)
(B) or PBS (500 ng/ml) (C) at designated time points and collected 3 hours after
treatment. n = 6 to 9 replicates from two to three independent experiments per
time point. Data were normalized to basal Chi3l1 expression in PBS control cells
(depicted by dashed line) (B) or expression at 18 hours (C). Main effect P = 0.0150
(B) and P = 0.0013 (C). Multiple comparison tests are depicted on graphs. (D) qPCR
showing LPS-induced Chi3l1 expression in WT primary astrocytes after transfection
with control (siSCR) or Bmal1 (siBmal1) siRNA, treated with LPS (500 ng/ml) at 24 or
36 hours after synchronization [as in (B)]. n = 3 replicates per genotype per time point.
(E) Diagram depicting hypothesis that induction of Chi3l1 expression (blue) is de-
pendent on the circadian phase of BMAL1 transcriptional activity (green curve).
(F) qPCR showing gene expression from WT primary astrocytes ± 10 M A fibrils
for 48 to 72 hours. n = 6 replicates from two independent experiments. All data
represent means ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001. Analyzed by two-way
ANOVA (A and D) or one-way ANOVA (B and C) with Tukey correction for multiple
comparisons or two-tailed Student’s t test (F).
could be neuroprotective in AD but destructive in settings of
acute inflammation.
In APP/PS1 mice, we observed that Chi3l1 deletion causes de-
creased periplaque astrocyte clustering. Attenuation of astrocyte
activation in APP/PS1 mice has previously yielded mixed results, as
Gfap:Vimentin double KO increases (33), whereas Stat3 KO decreases
Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020
(34) plaque burden. Our finding that Chi3l1 knockdown enhances
phagocytosis of zymosan beads and A by cultured astrocytes sug-
gests that Chi3l1/YKL-40 is a general regulator of astrocyte phago-
cytosis and reveals a potential mechanism for the decrease in
amyloid plaques in vivo. Attenuated astrocyte activation in APP/
PS1 models has previously been associated with increased mi-
croglial plaque clustering (33, 34) and, in one case, with increased
microglial phagocytosis of A (34), which is consistent with our data.
Decreased astrocyte activation in Chi3l1 KO mice (and in these other
models) could relent a physical barrier that previously limited
microglial access to the plaque. Alternatively, astrocyte-secreted
YKL-40 may signal to restrain microglial phagocytic activation.
The exacerbation of LPS-induced microglial transcriptional changes
with Chi3l1 deletion matches previous reports showing that Chi3l1
can cell-autonomously suppress the macrophage inflammatory
Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020
response in the periphery (39). Together, these data suggest that
Chi3l1 could also be important in microglia. Recently, a single-­
nucleus RNAseq study identified Chi3l1 as a gene that is strongly
up-regulated in microglia in the brains of human patients with AD
(41). Whereas our single-nucleus RNAseq data show clear expres-
sion of Chi3l1 in astrocytes, a small number of microglia do appear
to express Chi3l1, and this could increase in the setting of disease.
Thus, a cell-autonomous effect of Chi3l1 in microglia is highly pos-
sible. Our observation that Chi3l1 knockdown in cultured microglia
increases phagocytosis of beads and A suggests that the increase
in periplaque CD68 expression in Chi3l1 KO;APP/PS1 mice may be
due to a cell-autonomous effect in microglia. In this case, increased
Chi3l1 expression in microglia in AD [as reported by Zhou et al.
(41)] would be expected to suppress microglial, and potentially astro-
cytic, A phagocytosis and accelerate plaque growth, in keeping with
our data.
In patients with AD, we observed that a common variant in the
CHI3L1 gene that causes decreased CSF YKL-40 concentrations is
associated with slower AD progression. This observation was made
in a cohort of extremely well-characterized individuals confirmed
to have AD by both clinical evaluation and biomarkers (based on
CSF A/tau or amyloid positron emission tomography imaging).
Because CSF YKL-40 increases in response to aging, inflammation,
and neurodegeneration, it is difficult to determine how YKL-40
itself might be affecting these processes simply by measuring it in
CSF. By examining disease progression in carriers of this genetic
variant, which lowers CHI3L1/YKL-40 expression throughout life,
we can assume that changes in progression are likely caused by re-
duced CHI3L1/YKL-40 signaling, providing a unique opportunity
to assess a possible causal relationship between Chi3l1/YKL-40 expres-
sion and AD. Our results suggest that inhibition of Chi3l1/YKL-40
may be a potential future therapeutic target for limiting plaque
accumulation, optimizing the glial phagocytic response to plaques,
and slowing progression of AD.
Research into the relationship between circadian disruption and
neurodegenerative disease has begun to uncover a role for the clock
in regulating astrogliosis (24), microgliosis (22), and plaque deposi-
tion (17). Our findings provide an example of a clock-controlled gene
(Chi3l1) that does not oscillate at the mRNA level, likely because of
a long mRNA half-life. However, because Chi3l1 induction by LPS
is suppressed with Bmal1 deletion and is greatest at times when BMAL1
transcriptional activity is highest, it appears that BMAL1 transcrip-
tional activity gates the induction of Chi3l1 in astrocytes, revealing
a role for circadian timing in Chi3l1 regulation.
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As Chi3l1 is strongly suppressed in Bmal1 KO astrocytes, it is
important to reconcile the reduction in A plaque burden that we
have observed in Chi3l1 KO mice with our previous data showing
that global Bmal1 deletion increased fibrillar plaque burden (17).
Global Bmal1 deletion affects every cell type in the brain while also
disrupting peripheral clocks, sleep-wake cycles, and whole-animal
rhythmicity. The resultant phenotype in global Bmal1 KO mice is
thus a summation of many smaller and possibly divergent effects. It
is likely that in global Bmal1 KO mice, the potentially beneficial
effect of decreased Chi3l1 on plaques is overwhelmed by effects on
sleep and other processes, resulting in a net increase in plaques.
Astrocyte-specific effects of Bmal1 deletion on amyloid plaque depo-
sition remain to be explored. Elucidating the intricacies of competing
cell-specific pathways regulated by the clock in the brain is vital in
understanding how circadian dysfunction may affect the course of
neurodegeneration (42).
There are several limitations to this study. Our experiments do
not differentiate the relative effects of Chi3l1 in astrocytes versus
microglia in vivo. Such disentanglement would require cell type–
specific Chi3l1 manipulation. The APP/PS1 mouse used in this study
models amyloid plaque formation due to rare familial mutations and
does not recapitulate all aspects of human AD, especially tau aggrega-
tion. Future studies in alternate models, such as tau transgenic mice, will
be needed. The effects of Chi3l1 on phagocytosis of various A and
tau species, such as oligomers, were not investigated. Last, our clinical
data do not establish how CHI3L1 polymorphisms affect different
aspects of AD pathology or glial activation in humans. Human
pathological and biomarker data will need to be integrated with
CHI3L1 genotyping in future studies.
In summary, we have provided evidence that Chi3l1 regulates glial
activation, A phagocytosis, and amyloid plaque deposition in mice
and influences AD progression in humans. These findings identify
Chi3l1/YKL-40 as a potential therapeutic target for slowing disease
progression in AD and provide insights into regulation of neuro­
inflammation by the astrocyte circadian clock.
MATERIALS AND METHODS
Study design
The goal of this study was to elucidate the role of the well-known
biomarker of AD, Chi3l1/YKL-40, in neuroinflammation and AD
pathogenesis. We used data from a large observation study of AD to
determine whether a known genetic variant in the CHI3L1 gene in
humans, which was associated with lower CSF YKL-40 levels, might
influence the rate of AD progression. This analysis method was
based on previous work by members of our group in identifying the
variant in CHI3L1 that affects YKL-40 levels and in developing a
method to accurately detect single-gene influences on clinical AD
progression (26). Mouse studies were then carried out using consti-
tutive Chi3l1−/− mice, which were crossed to an APP/PS1 model of
-amyloidosis for some experiments. Last, the regulation of Chi3l1
expression by the circadian clock was investigated using a variety of
tissue-specific Bmal1 KO mice, as well as other circadian clock gene
mutant mice. Cell culture experiments using primary mouse glial
cultures and siRNA were also used for mechanistic studies. Sample
size for APP/PS1-21 experiments was determined at the outset and
based on power calculations derived from previous analysis of
amyloid plaque pathology in this line from our laboratory. Mice
were not randomized but were grouped on the basis of genotype
Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020
with groups containing roughly equal numbers of male and female
animals. Mouse tissue samples were all processed together and ana-
lyzed to prevent batch effects. Investigators were blinded to geno-
type throughout the data analysis process. All mice were housed in
the same facility, and cohorts of mice were bred at the same time
such that they aged together. The number of biological replicates is
indicated in the figure legends.
Mice
All mouse experiments were conducted in accordance with protocols
approved by the Washington University Institutional Animal Care and
Use Committee. Bmal1−/− and Aldh1L1-CreERT2+, CX3CR1-CreERT2+,
and Bmal1f/f mice were obtained from the Jackson laboratory and
bred at Washington University. Tissue from NPAS2 KO, CLOCK KO,
and CLOCK/NPAS2 double KO was provided by D. Weaver
Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020
(University of Massachusetts, Worcester, MA). Chi3l1−/− mice were
obtained from J. Elias (Brown University, Providence, RI). APP/
PS1-21 mice were obtained from M. Jucker (University of Tübingen,
Tübingen, Germany). Timed-pregnant WT CD1 mice for culture
experiments were obtained from Charles River Laboratories (Wilm-
ington, MA). All mice except those used for perinatal cultures were
maintained on a C57BL/6 background and housed under 12-hour
light/12-hour dark conditions, unless otherwise specified. All mice
expressing any Cre or APP/PS1 transgene were heterozygous for these
transgenes. Aldh1L1-CreERT2+;Bmal1f/f and CX3CR1-CreERT2+; Bmal1f/f
mice were given tamoxifen (dissolved in corn oil, 2 mg per mouse per
day for 5 days; Sigma-Aldrich) by oral gavage at 1 or 2 months, re-
spectively, to induce Bmal1 deletion in the applicable tissue. Cre−,
Bmal1f/f control littermates were given identical tamoxifen treatment.
Human studies
For human studies, data were collected as part of several ongoing
longitudinal observational studies of aging and dementia carried
out at the Knight Alzheimer’s Disease Research Center. Participants
are evaluated annually by clinical staff who are blinded to the partic-
ipant’s previous diagnosis and all previously collected data, allowing
an unbiased assessment of AD diagnosis CDR each year (43). The
inclusion/exclusion criteria for our analyses were predetermined,
and only data from participants meeting these criteria were queried.
Inclusion required a clinical AD diagnosis at the last visit, available
CSF biomarkers with a profile compatible with AD (based on CSF
A/tau profiles with established cutoffs), a CDR > 0 at last assess-
ment, and at least 1.5 years of follow-up. Exclusions included a
clinical diagnosis of a non-AD form of dementia or diagnoses of
another coexistent neurological diseases. In total, 778 participants
were included.
Statistical analysis
For mouse and cell experiments, statistical analyses were performed
using GraphPad Prism version 8.02. When multiple t tests were per-
formed, Holm-Sidak correction test for multiple comparisons was
applied unless otherwise noted. Fluidigm qPCR data were analyzed
by two-way [individual gene or gene groups provided no baseline
change in phosphate-buffered saline (PBS) or APP/PS1− mice] or
three-way [if a significant main effect of genotype was found by
two-way analysis of variance (ANOVA) at baseline in PBS or APP/
PS1− mice] ANOVA with Tukey correction for multiple compari-
sons. Other tests are noted in figure legends. In mouse and cell experi-
ments, data points were determined to be outliers (and thus excluded)
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based on the ROUT method in Prism 8, Q = 1%, performed post
hoc where appropriate.
Statistical analysis of human AD progression was carried out us-
ing R statistical software, and the package nlme was used for a linear
mixed model. A linear mixed-model repeated measure framework
was used to account for correlation between repeated measures in
the same individual. Disease progression was modeled as follows
(Y ) =  _ 1〖 [ SNP * time ] + 〗 _ 2 [〖CDR _ baseline8 * time ] + 〗 _ 3
​​ 〖age 
     _ baseline 〗 _ 4〖gender + 〗 _ 5 education  +   _ 6 SNP  + 
         _​  7​ ​​ S. A. Rosenberg, Y. Li, A. M. Nour, C. R. Parikh, I. Schmidt, Y. Modis, L. Cantley, J. A. Elias,
CDR _ (baseline  +  )  _ 8〖time + 〗 _ 9 PC1〖 + 〗 _ 10 PC2
where Y was CDR-SB, the change in CDR-SB per year baseline CDR,
baseline age, gender, follow-up time, level of education, and, to
avoid the possibility of spurious association due to population sub-
structure, the two first principal components scores were included
as covariates.
SUPPLEMENTARY MATERIALS
stm.sciencemag.org/cgi/content/full/12/574/eaax3519/DC1
Materials and Methods
Fig. S1. Loss of Chi3l1 exacerbates the LPS-induced inflammatory response.
Fig. S2. Loss of Chi3l1 does not change astrocyte reactivity gene signature but does modulate
the microglial response to LPS.
Fig. S3. Loss of Chi3l1 mitigates amyloid pathology.
Fig. S4. Plaque number and size reduced with Chi3l1 deletion without affecting
APP processing.
Fig. S5. Loss of Chi3l1 mitigates plaque-related astrogliosis.
Fig. S6. Loss of Chi3l1 alters plaque-related microglial activation.
Fig. S7. Loss of Chi3l1 reduces inflammation in APP/PS1+ mouse hippocampus.
Fig. S8. Chi3l1 is regulated by the circadian clock and expressed in astrocytes.
View/request a protocol for this paper from Bio-protocol.
REFERENCES AND NOTES
1. M. Colonna, O. Butovsky, Microglia function in the central nervous system during health
and neurodegeneration. Annu. Rev. Immunol. 35, 441–468 (2017).
2. A. M. Arranz, B. De Strooper, The role of astroglia in Alzheimer's disease: Pathophysiology
and clinical implications. Lancet Neurol. 18, 406–414 (2019).
3. S. A. Liddelow, K. A. Guttenplan, L. E. Clarke, F. C. Bennett, C. J. Bohlen, L. Schirmer,
M. L. Bennett, A. E. Münch, W.-S. Chung, T. C. Peterson, D. K. Wilton, A. Frouin, B. A. Napier,
N. Panicker, M. Kumar, M. S. Buckwalter, D. H. Rowitch, V. L. Dawson, T. M. Dawson,
B. Stevens, B. A. Barres, Neurotoxic reactive astrocytes are induced by activated
microglia. Nature 541, 481–487 (2017).
4. R. Craig-Schapiro, R. J. Perrin, C. M. Roe, C. Xiong, D. Carter, N. J. Cairns, M. A. Mintun,
E. R. Peskind, G. Li, D. R. Galasko, C. M. Clark, J. F. Quinn, G. D'Angelo, J. P. Malone,
R. R. Townsend, J. C. Morris, A. M. Fagan, D. M. Holtzman, YKL-40: A novel prognostic fluid
biomarker for preclinical Alzheimer's disease. Biol. Psychiatry 68, 903–912 (2010).
5. C. L. Sutphen, M. S. Jasielec, A. R. Shah, E. M. Macy, C. Xiong, A. G. Vlassenko,
T. L. S. Benzinger, E. E. J. Stoops, H. M. J. Vanderstichele, B. Brix, H. D. Darby,
M. L. J. Vandijck, J. H. Ladenson, J. C. Morris, D. M. Holtzman, A. M. Fagan, Longitudinal
cerebrospinal fluid biomarker changes in preclinical Alzheimer disease during middle
age. JAMA Neurol. 72, 1029–1042 (2015).
6. F. Llorens, K. Thüne, W. Tahir, E. Kanata, D. Diaz-Lucena, K. Xanthopoulos, E. Kovatsi,
C. Pleschka, P. Garcia-Esparcia, M. Schmitz, D. Ozbay, S. Correia, Â. Correia, I. Milosevic,
O. Andréoletti, N. Fernández-Borges, I. M. Vorberg, M. Glatzel, T. Sklaviadis, J. M. Torres,
S. Krasemann, R. Sánchez-Valle, I. Ferrer, I. Zerr, YKL-40 in the brain and cerebrospinal
fluid of neurodegenerative dementias. Mol. Neurodegener. 12, 83 (2017).
7. C. Malmeström, M. Axelsson, J. Lycke, H. Zetterberg, K. Blennow, B. Olsson, CSF levels
of YKL-40 are increased in MS and replaces with immunosuppressive treatment.
J. Neuroimmunol. 269, 87–89 (2014).
8. I. Illán-Gala, D. Alcolea, V. Montal, O. Dols-Icardo, L. Muñoz, N. de Luna, J. Turón-Sans,
E. Cortés-Vicente, M. B. Sánchez-Saudinós, A. Subirana, I. Sala, R. Blesa, J. Clarimón,
J. Fortea, R. Rojas-Garcia, A. Lleó, CSF sAPP, YKL-40, and NfL along the ALS-FTD
spectrum. Neurology 91, e1619–e1628 (2018).
Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020
9. D. Alcolea, E. Vilaplana, M. Suárez-Calvet, I. Illán-Gala, R. Blesa, J. Clarimón, A. Lladó,
R. Sánchez-Valle, J. L. Molinuevo, G. García-Ribas, Y. Compta, M. J. Martí, G. Piñol-Ripoll,
G. Amer-Ferrer, A. Noguera, A. García-Martín, J. Fortea, A. Lleó, CSF sAPP, YKL-40,
and neurofilament light in frontotemporal lobar degeneration. Neurology 89, 178–188
(2017).
10. M. Querol-Vilaseca, M. Colom-Cadena, J. Pegueroles, C. San Martín-Paniello, J. Clarimon,
O. Belbin, J. Fortea, A. Lleó, YKL-40 (chitinase 3-like I) is expressed in a subset of astrocytes
in Alzheimer's disease and other tauopathies. J. Neuroinflammation 14, 118 (2017).
11. R. Bhardwaj, J. W. Yester, S. K. Singh, D. D. Biswas, M. J. Surace, M. R. Waters, K. F. Hauser,
Z. Yao, B. F. Boyce, T. Kordula, RelB/p50 complexes regulate cytokine-induced YKL-40
expression. J. Immunol. 194, 2862–2870 (2015).
12. C. H. He, C. G. Lee, C. S. Dela Cruz, C.-M. Lee, Y. Zhou, F. Ahangari, B. Ma, E. L. Herzog,
Chitinase 3-like 1 regulates cellular and tissue responses via IL-13 receptor 2. Cell Rep. 4,
830–841 (2013).
13. D. Bonneh-Barkay, G. Wang, W. A. Laframboise, C. A. Wiley, S. J. Bissel, Exacerbation
of experimental autoimmune encephalomyelitis in the absence of breast regression
protein 39/chitinase 3-like 1. J. Neuropathol. Exp. Neurol. 71, 948–958 (2012).
Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020
14. C. A. Wiley, D. Bonneh-Barkay, C. E. Dixon, A. Lesniak, G. Wang, S. J. Bissel, P. M. Kochanek,
Role for mammalian chitinase 3-like protein 1 in traumatic brain injury. Neuropathology
35, 95–106 (2015).
15. E. S. Musiek, M. Bhimasani, M. A. Zangrilli, J. C. Morris, D. M. Holtzman, Y.-E. S. Ju, Circadian
rest-activity pattern changes in aging and preclinical Alzheimer disease. JAMA Neurol. 75,
582–590 (2018).
16. E. S. Musiek, D. M. Holtzman, Mechanisms linking circadian clocks, sleep,
and neurodegeneration. Science 354, 1004–1008 (2016).
17. G. J. Kress, F. Liao, J. Dimitry, M. R. Cedeno, G. A. FitzGerald, D. M. Holtzman, E. S. Musiek,
Regulation of amyloid- dynamics and pathology by the circadian clock. J. Exp. Med. 215,
1059–1068 (2018).
18. J. A. Mohawk, C. B. Green, J. S. Takahashi, Central and peripheral circadian clocks
in mammals. Annu. Rev. Neurosci. 35, 445–462 (2012).
19. A. M. Curtis, M. M. Bellet, P. Sassone-Corsi, L. A. O'Neill, Circadian clock proteins
and immunity. Immunity 40, 178–186 (2014).
20. L. M. Prolo, J. S. Takahashi, E. D. Herzog, Circadian rhythm generation and entrainment
in astrocytes. J. Neurosci. 25, 404–408 (2005).
21. L. K. Fonken, M. G. Frank, M. M. Kitt, R. M. Barrientos, L. R. Watkins, S. F. Maier, Microglia
inflammatory responses are controlled by an intrinsic circadian clock. Brain Behav.
Immun. 45, 171–179 (2015).
22. R. Nakazato, S. Hotta, D. Yamada, M. Kou, S. Nakamura, Y. Takahata, H. Tei, R. Numano,
A. Hida, S. Shimba, M. Mieda, E. Hinoi, Y. Yoneda, T. Takarada, The intrinsic microglial
clock system regulates interleukin-6 expression. Glia 65, 198–208 (2017).
23. E. S. Musiek, M. M. Lim, G. Yang, A. Q. Bauer, L. Qi, Y. Lee, J. H. Roh, X. Ortiz-Gonzalez,
J. T. Dearborn, J. P. Culver, E. D. Herzog, J. B. Hogenesch, D. F. Wozniak, K. Dikranian,
B. I. Giasson, D. R. Weaver, D. M. Holtzman, G. A. Fitzgerald, Circadian clock proteins
regulate neuronal redox homeostasis and neurodegeneration. J. Clin. Invest. 123,
5389–5400 (2013).
24. B. V. Lananna, C. J. Nadarajah, M. Izumo, M. R. Cedeño, D. D. Xiong, J. Dimitry, C. F. Tso,
C. A. McKee, P. Griffin, P. W. Sheehan, J. A. Haspel, B. A. Barres, S. A. Liddelow,
J. S. Takahashi, I. N. Karatsoreos, E. S. Musiek, Cell-autonomous regulation of astrocyte
activation by the circadian clock protein BMAL1. Cell Rep. 25, 1–9.e5 (2018).
25. J. L. Del-Aguila, Z. Li, U. Dube, K. A. Mihindukulasuriya, J. P. Budde, M. V. Fernandez,
L. Ibanez, J. Bradley, F. Wang, K. Bergmann, R. Davenport, J. C. Morris, D. M. Holtzman,
R. J. Perrin, B. A. Benitez, J. Dougherty, C. Cruchaga, O. Harari, A single-nuclei RNA
sequencing study of Mendelian and sporadic AD in the human brain. Alzheimers Res. Ther.
11, 71 (2019).
26. Y. Deming, K. Black, D. Carrell, Y. Cai, J. L. Del-Aguila, M. V. Fernandez, J. Budde, S. Ma,
B. Saef, B. Howells, S. Bertelsen, K.-l. Huang, C. L. Sutphen, R. Tarawneh, A. M. Fagan,
D. M. Holtzman, J. C. Morris, A. M. Goate, J. D. Dougherty, C. Cruchaga, Chitinase-3-like 1
protein (CHI3L1) locus influences cerebrospinal fluid levels of YKL-40. BMC Neurol. 16, 217
(2016).
27. J. L. Del-Aguila, M. V. Fernández, S. Schindler, L. Ibanez, Y. Deming, S. Ma, B. Saef, K. Black,
J. Budde, J. Norton, R. Chasse; Alzheimer’s Disease Neuroimaging Initiative (ADNI),
O. Harari, A. Goate, C. Xiong, J. C. Morris, C. Cruchaga, Assessment of the genetic
architecture of Alzheimer's disease risk in rate of memory decline. J. Alzheimers Dis. 62,
745–756 (2018).
28. Y. Zhang, S. A. Sloan, L. E. Clarke, C. Caneda, C. A. Plaza, P. D. Blumenthal, H. Vogel,
G. K. Steinberg, M. S. B. Edwards, G. Li, J. A. Duncan III, S. H. Cheshier, L. M. Shuer,
E. F. Chang, G. A. Grant, M. G. H. Gephart, B. A. Barres, Purification and characterization
of progenitor and mature human astrocytes reveals transcriptional and functional
differences with mouse. Neuron 89, 37–53 (2016).
29. Y. Zhang, K. Chen, S. A. Sloan, M. L. Bennett, A. R. Scholze, S. O'Keeffe, H. P. Phatnani,
P. Guarnieri, C. Caneda, N. Ruderisch, S. Deng, S. A. Liddelow, C. Zhang, R. Daneman,
11 of 12
SCIENCE TRANSLATIONAL MEDICINE | RESEARCH ARTICLE
T. Maniatis, B. A. Barres, J. Q. Wu, An RNA-sequencing transcriptome and splicing
database of glia, neurons, and vascular cells of the cerebral cortex. J. Neurosci. 34,
11929–11947 (2014).
30. C. G. Lee, D. Hartl, G. R. Lee, B. Koller, H. Matsuura, C. A. Da Silva, M. H. Sohn, L. Cohn,
R. J. Homer, A. A. Kozhich, A. Humbles, J. Kearley, A. Coyle, G. Chupp, J. Reed, R. A. Flavell,
J. A. Elias, Role of breast regression protein 39 (BRP-39)/chitinase 3-like-1 in Th2
and IL-13-induced tissue responses and apoptosis. J. Exp. Med. 206, 1149–1166 (2009).
31. R. Radde, T. Bolmont, S. A. Kaeser, J. Coomaraswamy, D. Lindau, L. Stoltze, M. E. Calhoun,
F. Jäggi, H. Wolburg, S. Gengler, C. Haass, B. Ghetti, C. Czech, C. Hölscher, P. M. Mathews,
M. Jucker, A42-driven cerebral amyloidosis in transgenic mice reveals early and robust
pathology. EMBO Rep. 7, 940–946 (2006).
32. S. D. Styren, R. L. Hamilton, G. C. Styren, W. E. Klunk, X-34, a fluorescent derivative
of Congo red: A novel histochemical stain for Alzheimer's disease pathology.
J. Histochem. Cytochem. 48, 1223–1232 (2000).
33. A. W. Kraft, X. Hu, H. Yoon, P. Yan, Q. Xiao, Y. Wang, S. C. Gil, J. Brown, U. Wilhelmsson,
J. L. Restivo, J. R. Cirrito, D. M. Holtzman, J. Kim, M. Pekny, J.-M. Lee, Attenuating astrocyte
activation accelerates plaque pathogenesis in APP/PS1 mice. FASEB J. 27, 187–198 (2013).
34. N. Reichenbach, A. Delekate, M. Plescher, F. Schmitt, S. Krauss, N. Blank, A. Halle,
G. C. Petzold, Inhibition of Stat3-mediated astrogliosis ameliorates pathology
in an Alzheimer's disease model. EMBO Mol. Med. 11, e9665 (2019).
35. M. Koistinaho, S. Lin, X. Wu, M. Esterman, D. Koger, J. Hanson, R. Higgs, F. Liu, S. Malkani,
K. R. Bales, S. M. Paul, Apolipoprotein E promotes astrocyte colocalization
and degradation of deposited amyloid- peptides. Nat. Med. 10, 719–726 (2004).
36. J. L. Zamanian, L. Xu, L. C. Foo, N. Nouri, L. Zhou, R. G. Giffard, B. A. Barres, Genomic
analysis of reactive astrogliosis. J. Neurosci. 32, 6391–6410 (2012).
37. N. Koike, S.-H. Yoo, H.-C. Huang, V. Kumar, C. Lee, T.-K. Kim, J. S. Takahashi, Transcriptional
architecture and chromatin landscape of the core circadian clock in mammals. Science
338, 349–354 (2012).
38. S. Janelidze, N. Mattsson, E. Stomrud, O. Lindberg, S. Palmqvist, H. Zetterberg,
K. Blennow, O. Hansson, CSF biomarkers of neuroinflammation and cerebrovascular
dysfunction in early Alzheimer disease. Neurology 91, e867–e877 (2018).
39. C. S. Dela Cruz, W. Liu, C. H. He, A. Jacoby, A. Gornitzky, B. Ma, R. Flavell, C. G. Lee,
J. A. Elias, Chitinase 3-like-1 promotes Streptococcus pneumoniae killing and augments
host tolerance to lung antibacterial responses. Cell Host Microbe 12, 34–46 (2012).
40. J. Y. Choi, I. J. Yeo, K. C. Kim, W. R. Choi, J.-K. Jung, S.-B. Han, J. T. Hong, K284-6111
prevents the amyloid beta-induced neuroinflammation and impairment of recognition
memory through inhibition of NF-B-mediated CHI3L1 expression. J. Neuroinflammation
15, 224 (2018).
41. Y. Zhou, W. M. Song, P. S. Andhey, A. Swain, T. Levy, K. R. Miller, P. L. Poliani, M. Cominelli,
S. Grover, S. Gilfillan, M. Cella, T. K. Ulland, K. Zaitsev, A. Miyashita, T. Ikeuchi, M. Sainouchi,
Lananna et al., Sci. Transl. Med. 12, eaax3519 (2020) 16 December 2020
A. Kakita, D. A. Bennett, J. A. Schneider, M. R. Nichols, S. A. Beausoleil, J. D. Ulrich,
D. M. Holtzman, M. N. Artyomov, M. Colonna, Human and mouse single-nucleus
transcriptomics reveal TREM2-dependent and TREM2-independent cellular responses
in Alzheimer's disease. Nat. Med. 26, 131–142 (2020).
42. B. V. Lananna, E. S. Musiek, The wrinkling of time: Aging, inflammation, oxidative stress,
and the circadian clock in neurodegeneration. Neurobiol. Dis. 139, 104832 (2020).
43. J. C. Morris, S. Weintraub, H. C. Chui, J. Cummings, C. DeCarli, S. Ferris, N. L. Foster,
D. Galasko, N. Graff-Radford, E. R. Peskind, D. Beekly, E. M. Ramos, W. A. Kukull, The
uniform data set (UDS): Clinical and cognitive variables and descriptive data
from Alzheimer disease centers. Alzheimer Dis. Assoc. Disord. 20, 210–216 (2006).
Acknowledgments: We thank D. Weaver (UMass Medical School) for Clock and Npas2 KO
mouse tissue and the Washington University Center for Cellular Imaging (WUCCI) for imaging
assistance. Funding: This work was supported by NIH grants R01AG054517 (E.S.M) and
R01AG044546, P01AG003991, RF1AG053303, R01AG058501, U01AG058922, U01AG052411,
and R01AG05777 (C.C.). WUCCI is supported by the Washington University School of Medicine,
The Children’s Discovery Institute of WU and St. Louis Children’s Hospital (CDI-CORE-2015-505),
and the Foundation for Barnes-Jewish Hospital (3770). The Knight ADRC at Washington
Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020
University is funded by NIA grants P50-AG05681, P01-AG03991, and P01-AG026276. Author
contributions: B.V.L. and E.S.M. designed and conducted mouse and cell experiments,
analyzed data, and wrote the manuscript. M.W.K. conducted and analyzed phagocytosis
experiments. C.A.M., J.M.D, C.J.N., D.D.X., and A.J.C conducted mouse and cell experiments.
C.G. and J.Z. performed and analyzed ChIP-qPCR. J.L.D.-A., F.H.G.F., and C.C. analyzed human
genetic data and contributed to the manuscript. J.A.E. provided Chi3l1 KO mice. Competing
interests: E.S.M. has consulted for Eisai Pharmaceuticals. No patents related to this work are
pending. The other authors declare that they have no competing interest. Data and materials
availability: All the data associated with this study are present in the paper or the
Supplementary Materials. Microarray data are freely available on the EMBL-EBI ArrayExpress
database, accession no. E-MTAB-7151.
Submitted 15 March 2019
Resubmitted 21 April 2020
Accepted 29 August 2020
Published 16 December 2020
10.1126/scitranslmed.aax3519
Citation: B. V. Lananna, C. A. McKee, M. W. King, J. L. Del-Aguila, J. M. Dimitry, F. H. G. Farias,
C. J. Nadarajah, D. D. Xiong, C. Guo, A. J. Cammack, J. A. Elias, J. Zhang, C. Cruchaga, E. S. Musiek,
Chi3l1/YKL-40 is controlled by the astrocyte circadian clock and regulates neuroinflammation
and Alzheimer’s disease pathogenesis. Sci. Transl. Med. 12, eaax3519 (2020).
12 of 12
Chi3l1/YKL-40 is controlled by the astrocyte circadian clock and regulates
neuroinflammation and Alzheimer's disease pathogenesis
Brian V. Lananna, Celia A. McKee, Melvin W. King, Jorge L. Del-Aguila, Julie M. Dimitry, Fabiana H. G. Farias, Collin J.
Nadarajah, David D. Xiong, Chun Guo, Alexander J. Cammack, Jack A. Elias, Jinsong Zhang, Carlos Cruchaga and Erik
S. Musiek

Sci Transl Med 12, eaax3519.

DOI: 10.1126/scitranslmed.aax3519

Downloaded from http://stm.sciencemag.org/ at GOTEBORGS UNIV on December 16, 2020

Targeting astrocytes in AD
Astrocytes and microglia play a dual role in Alzheimer's disease (AD), increasing neuroinflammation and
limiting plaque growth through phagocytic activity. The astrocytic protein YKL-40 is increased in the cerebrospinal
fluid (CSF) of patients with AD; however, its role in AD pathophysiology is unclear. Here, Lananna et al. used
mouse model and in vitro systems to show that Chi3l1, the gene coding for YKL-40, plays a detrimental role in
AD. Its deletion reduced amyloid plaque formation and promoted A β phagocytosis. A polymorphism in CHI3L1
resulting in reduced protein expression was associated with slower AD progression in patients. The results
suggest that YKL-40 could be targeted for reducing AD progression in patients.

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