of 10
Articles
https://doi.org/10.1038/s41593-020-0589-7
Gut-seeded
α
-synuclein fibrils promote gut
dysfunction and brain pathology specifically
in aged mice
Collin Challis
1
, Acacia Hori
1,4
, Timothy R. Sampson
1,2,4
, Bryan B. Yoo
1
, Rosemary C. Challis
1
,
Adam M. Hamilton
2
, Sarkis K. Mazmanian
1
, Laura A. Volpicelli-Daley
3
and Viviana Gradinaru
1
*
1
Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA, USA.
2
Department of Physiology, Emory University School
of Medicine, Atlanta, GA, USA.
3
Center for Neurodegeneration and Experimental Therapeutics, University of Alabama at Birmingham, Birmingham, AL,
USA.
4
These authors contributed equally: Acacia Hori, Timothy R. Sampson. *e-mail: viviana@caltech.edu
SUPPLEMENTARY INFORMATION
In the format provided by the authors and unedited.
NATuRe NeuR
oSCieNCe
|
www.nature.com/natureneuroscience
Supplementary Table 1. List of antibodies used
Primary
Antibodies
Manufacturer
Cat. #
Usage notes
Mouse IgG2a monoclonal anti-α-synuclein phospho
(Ser129) (Clone81A)
BioLegend
MMS-5091
Duodenum IHC 1:300
Brain IHC 1:500
Rabbit polyclonal anti-α-synuclein phosphor (Ser129)
Abcam
ab59264
Western 1:500
Rabbit monoclonal anti-Alpha-synuclein filament antibody
[MJFR-14-6-4-2] - Conformation-Specific
Abcam
ab209538
Dot blot 2ng/mL
Rabbit polyclonal anti-Protein gene product 9.5 (PGP9.5) Millipore
AB1761-I
Duodenum IHC 1:300
Nodose IHC 1:100
Chicken polyclonal anti-Protein gene product 9.5 (PGP9.5) ThermoFisher Scientific PA1-10011
Duodenum IHC 1:300
Chicken polyclonal anti-Glial fibrillary acidic protein (GFAP) Millipore
AB5541
Duodenum IHC 1:300
Goat polyclonal anti-Choline acetyltransferase (ChAT)
Millipore
AB144P
Brain IHC 1:500
Rabbit polyclonal anti-Tyrosine hydroxylase (TH)
Millipore
AB152
Brain IHC 1:500
Chicken polyclonal anti-Green fluorescent protein (GFP) Aves Labs
GFP-1010
Brain IHC 1:1000
Rabbit polyclonal anti-Red fluorescent protein (RFP)
Rockland
600-401-379
Brain IHC 1:1000
Rabbit polyclonal anti-GBA1
Abcam
ab175869
Western 1:1000
Rabbit polyclonal anti-Interleukin 6 (IL6)
Abcam
ab7737
Western 1:500
Rabbit polyclonal anti-Iba1
Wako
016-20001
Western 1:1000
Rabbit polyclonal anti-β-Tubulin
Abcam
ab6046
Western 1:1000
Mouse IgG2b anti-β-actin
Cell Signaling
3700
Western 1:1000
Secondary Antibodies
AlexaFluor 488 Goat anti-Mouse IgG2a
ThermoFisher Scientific A-21131
IHC 1:300-1000
AlexaFluor 488 Donkey anti-Mouse IgG
ThermoFisher Scientific A-21202
IHC 1:300-1000
AlexaFluor 488 Donkey anti-Chicken IgY
Jackson
ImmunoResearch
703-545-155
IHC 1:300-1000
AlexaFluor 555 Donkey anti-Rabbit IgG
ThermoFisher Scientific A-31572
IHC 1:300-1000
AlexaFluor 555 Donkey anti-Goat IgG
ThermoFisher Scientific A-21432
IHC 1:300-1000
AlexaFluor 633 Goat anti-Rabbit IgG
ThermoFisher Scientific A-21071
IHC 1:300-1000
AlexaFluor 633 Goat anti-Chicken IgY
ThermoFisher Scientific A-21103
IHC 1:300-1000
Horseradish peroxidase (HRP)-linked Goat anti-Rabbit IgG CellSignaling
7074
Western, dot blot 1:2000
Horseradish peroxidase (HRP)-linked Goat anti-Mouse IgG CellSignaling
7076
Western 1:2000
Horseradish peroxidase (HRP)-linked Goat anti-Mouse
IgG2a
Abcam
ab97245
Western 1:2000
Supplementary Table 2. Statistics and quantification
Main figures
Figure
Number of subjects
Test
Summary
Key comparisons
1b
For 0, 7, 21, 60, 120 dpi:
α-Syn mon = 9, 9, 9, 8, 8
α-Syn PFF = 16, 14, 14, 11, 8
Two-way ANOVA
Condition x Time:
F(4,96) = 1.857; P = 0.1243
Time factor:
F(4,96) = 4.490; P = 0.0023
Condition factor:
F(1,96) = 22.72; P < 0.0001
PFF: 0 dpi vs. 21 dpi, ** p = 0.00
PFF: 0 dpi vs. 60 dpi, ** p = 0.0027
60 dpi: α-Syn PFF vs. α-Syn mon, * p
=0.0187
1c
Two-way ANOVA
Condition x Time:
F(4,96) = 3.980; P = 0.0050
Time factor:
F(4,96) = 1.808; P = 0.1136
Condition factor:
F(1,96) = 6.438; P = 0.0128
PFF: 0 dpi vs. 120 dpi, p = 0.0745
60 dpi: α-Syn PFF vs. α-Syn mon, * p
=0.0259
1d
For 0, 7, 21, 60, 120 dpi:
α-Syn mon = 6, 7, 6, 6, 6
α-Syn PFF = 7, 8, 9, 9, 8
Two-way ANOVA
Condition x Time:
F(4,62) = 5.264; P = 0.0010
Time factor:
F(4,62) = 3.042; P = 0.0235
Condition factor:
F(1,62) = 14.47; P = 0.0003
PFF: 0 dpi vs. 120 dpi, ** p = 0.0042
60 dpi: α-Syn PFF vs. α-Syn mon, ** p =
0.0012
120 dpi: α-Syn PFF vs. α-Syn mon, * p =
0.0239
1e
α-Syn mon = 4
α-Syn PFF = 4
Student’s t-test,
one-tailed
Fractalkine: t = 6.976, df = 3
IL-1a: t = 5.172, df = 3
IL-6: t = 2.562, df = 3
IL-7: t = 2.606, df = 3
MCP-1: t = 2.564, df = 3
MCSF: t = 2.430, df = 3
MIG: t = 2.572, df = 3
TECK: t = 2.711, df = 3
TIMP-2: t = 2.608, df = 3
Fractalkine, ** p = 0.0030
IL-1a, ** p = 0.0070
IL-6, * p = 0.0415
IL-7, * p = 0.0400
MCP-1, * p = 0.0416
MCSF, * p = 0.0467
MIG, * p = 0.0412
TECK, * p = 0.0365
TIMP-2, * p = 0.0400
1f
All conditions = 4
One-way ANOVA F(7,24) = 29.13, P < 0.0001
WT vs. PFF 21 dpi, * p = 0.0366
WT vs. PFF 60 dpi, **** p < 0.0001
WT vs. PFF 120 dpi, * p = 0.0245
WT vs. ASO, **** p < 0.0001
PFF 60 dpi vs. mon. 60 dpi, *** p = 0.0001
1h
WT = 6
ASO = 6
For 7, 21, 60, 120 dpi:
α-Syn PFF = 6, 6, 5, 5
For 7, 60 dpi:
α-Syn mon = 5, 5
One-way ANOVA F(8,45) = 1.519, P = 0.1776
WT vs. PFF 7dpi, * p = 0.0425
1i
One-way ANOVA F(8,45) = 2.501, P = 0.0269
WT vs. PFF 60 dpi, * p = 0.0329
WT vs. PFF 120 dpi, * p =0.0232
1k
All conditions = 4
One-way ANOVA F(8,27) = 6.622, P < 0.0001
WT vs. PFF 21 dpi, ** p = 0.0039
WT vs. PFF 60 dpi, * p = 0.0149
WT vs. PFF 120 dpi, * p = 0.0467
WT vs. ASO, * p = 0.0182
2b
All conditions = 4
One-way ANOVA F(8,27) = 3.230, P = 0.0116
WT vs. PFF 60 dpi, ** p = 0.0026
WT vs. PFF 120 dpi, * p = 0.0155
WT vs. ASO, ** p = 0.0032
2d
All conditions = 4
Except:
WT = 6, ASO = 5
One-way ANOVA F(8, 30) = 4.993, P = 0.0005
WT vs. PFF 60 dpi, ** p = 0.0042
WT vs. ASO, ** p = 0.0057
2e
WT = 11
ASO = 6
For 7, 21, 60, 120 dpi:
α-Syn PFF = 6, 5, 4, 4
For 7, 60 dpi:
α-Syn mon = 4, 4
One-way ANOVA F(8,40) = 4.697, P = 0.0004
WT vs. PFF 7dpi, * p = 0.0202
PFF 7 dpi vs. PFF 120 dpi, * p = 0.0415
WT vs. ASO, ** p = 0.0035
ASO vs. PFF 120 dpi, * p = 0.0109
2i
All conditions = 5
Two-way ANOVA
Genotype x Treatment
F(1,16) = 1.272; P = 0.2761
Genotype factor:
F(1,16) = 10.63; P = 0.0049
Treatment factor:
F(1,16) = 66.58; P < 0.0001
WT/GFP vs. WT/GBA1, **** p < 0.0001
WT/GFP vs. ASO/GBA1, * p = 0.0192
ASO/GFP vs. ASO/GBA1, *** p = 0.0008
WT/GBA1 vs. ASO/GBA1, * p = 0.0410
2j
All conditions = 5
Two-way ANOVA
Genotype x Treatment
F(1,16) = 3.685; P = 0.0729
Genotype factor:
F(1,16) = 49.90; P < 0.0001
Treatment factor:
F(1,16) = 3.801; P = 0.0690
WT/GFP vs. ASO/GFP, **** p < 0.0001
WT/GFP vs. ASO/GBA1, * p = 0.0139
ASO/GFP vs. ASO/GBA1, p = 0.0879
2k
For 0, 7, 21, 60 dpi:
WT = 17, 12, 11, 8
ASO = 13, 12, 11, 10
Fecal pellets:
Two-way ANOVA
Genotype x Time
F(3,86) = 1.160; P = 0.3296
Genotype factor:
F(1,86) = 13.80; P = 0.0004
Time factor:
F(3,86) = 2.084; P = 0.1082
0 dpi: WT vs. ASO, ** p = 0.0078
Pellet weight:
Two-way ANOVA
Genotype x Time
F(3,85) = 1.612; P = 0.1926
Genotype factor:
F(1,85) = 14.14; P = 0.0004
Time factor:
F(3,85) = 2.084; P = 0.1082
0 dpi: WT vs. ASO, ** p = 0.0061
2k
For 0, 7, 21, 60 dpi:
WT = 17, 12, 11, 8
ASO = 13, 12, 11, 10
Proportion water
weight:
Two-way ANOVA
Genotype x Time
F(3,85) = 6.410; P = 0.0006
Genotype factor:
F(1,85) = 54.34; P < 0.0001
Time factor:
F(3,85) = 0.6193; P = 0.6044
0 dpi: WT vs. ASO, **** p < 0.0001
7 dpi: WT vs. ASO, *** p = 0.0003
ASO: 0 dpi vs. 60 dpi, * p = 0.0265
Whole gut transit
time:
Two-way ANOVA
Genotype x Time
F(3,86) = 0.7794; P = 0.5087
Genotype factor:
F (1, 86) = 19.15; P < 0.0001
Time factor:
F(3,86) = 0.4554; P = 0.7142
0 dpi: WT vs. ASO, * p = 0.0123
3i
jR+
Baseline = 2(63)
WT = 3(183)
PFF 7 dpi = 4(355)
PFF 60 dpi = 3(110)
Mon 7 dpi = 3(110)
Mon 60 dpi = 3(90)
One-way ANOVA
Peak ΔF/F
F (5, 949) = 60.52; P < 0.0001
WT, stim vs. no stim, **** p < 0.0001
WT vs. PFF 7 dpi, **** p < 0.0001
WT vs. PFF 60 dpi, **** p < 0.0001
Area under the curve
F(5,932) = 55.90; P < 0.0001
WT, stim vs. no stim, **** p < 0.0001
WT vs. PFF 7 dpi, **** p < 0.0001
WT vs. PFF 60 dpi, **** p < 0.0001
3i
jR/ChR
Baseline = 2(12)
WT = 3(28)
PFF 7 dpi = 4(55)
PFF 60 dpi = 3(40)
Mon 7 dpi = 3(23)
Mon 60 dpi = 3(15)
One-way ANOVA
Peak ΔF/F
F (5, 157) = 23.49; P < 0.0001
WT, stim vs. no stim, **** p < 0.0001
WT vs. PFF 7 dpi, **** p < 0.0001
WT vs. PFF 60 dpi, **** p < 0.0001
Area under the curve
F(5,167) = 8.613; P < 0.0001
WT, stim vs. no stim, **** p < 0.001
WT vs. PFF 7 dpi, * p = 0.0488
WT vs. PFF 60 dpi, * p = 0.0214
3j
jR+
WT 0 dpvi = 3(131)
WT 7 dpvi = 3(78)
WT 60 dpvi = 3(131)
ASO 0 dpvi = 3(78)
ASO 7 dpvi = 3(80)
ASO 60 dpvi = 3(71)
Two-way ANOVA
Peak ΔF/F
Genotype x Time
F(2,563) = 10.01; P < 0.0001
Genotype factor
F (1, 563) = 68.91
Time factor
F(2,563) = 3.847; P = 0.0219
WT vs. ASO, **** p < 0.0001
WT vs. ASO 60 dpvi, * p = 0.0296
ASO vs. ASO 60 dpvi, *** p = 0.0007
ASO 7 dpvi vs. ASO 60 dpvi, ** p = 0.0068
Area under the curve
Genotype x Time
F(2,72) = 1.407; P = 0.2516
Genotype factor
F(1,72) = 48.71; P < 0.0001
Time factor
F (2, 72) = 0.8416; P = 0.4352
WT vs. ASO, **** p < 0.0001
WT vs. ASO 60 dpvi, *** p = 0.0006
WT 7 dpvi vs. ASO 7 dpvi, ** p = 0.0031
ASO vs. ASO 60 dpvi, p = 0.0808
3j
jR/ChR
WT 0 dpvi = 3(131)
WT 7 dpvi = 3(78)
WT 60 dpvi = 3(131)
ASO 0 dpvi = 3(78)
ASO 7 dpvi = 3(80)
ASO 60 dpvi = 3(71)
Two-way ANOVA
Peak ΔF/F
Genotype x Time
F (2, 490) = 7.103; P = 0.0009
Genotype factor
F (1, 490) = 61.42; P < 0.0001
Time factor
F (2, 490) = 1.595; P = 0.2040
0 dpi: WT vs. ASO, **** p < 0.0001
7 dpi: WT vs. ASO, *** p = 0.0006
Area under the curve
Genotype x Time
F (2, 72) = 1.586; P = 0.2117
Genotype factor
F (1, 72) = 10.55; P = 0.0018
Time factor
F (2, 72) = 3.074; P = 0.0523
WT vs. ASO, * p = 0.0398
4a
WT = 11
Aged = 5
ASO = 6
One-way ANOVA F(2,19) = 24.13; P < 0.0001
WT vs Aged, ** p = 0.0022
WT vs. ASO, **** p < 0.0001
4b
WT = 6
Aged = 4
ASO = 5
One-way ANOVA F(2,12) = 14.09; P = 0.0007
WT vs. ASO, *** p = 0.0006
Aged vs. ASO, * p = 0.0202
4c
WT = 4
Aged = 4
ASO = 6
One-way ANOVA F(2,11) = 4.298; P = 0.0418
WT vs. ASO, * p = 0.0468
4d
For 0, 60, 120 dpi:
α-Syn mon. = 9, 6, 4
α-Syn PFF. = 10, 7, 6
Two-way ANOVA
Treatment x Time
F(2,36) = 2.635; P = 0.0855
Time factor
F(2,36) = 9.165; P = 0.0006
Treatment factor
F(1,36) = 2.148; P = 0.1514
PFF: 0 dpi vs. 60 dpi, * p = 0.0300
PFF: 0 dpi vs. 120 dpi, *** p = 0.0007
4e
Two-way ANOVA
Treatment x Time
F(2,36) = 3.713; P = 0.0342
Time factor
F(2,36) = 2.426; P = 0.1027
Treatment factor
F(1,36) = 18.63; P = 0.0001
PFF: 0 dpi vs. 120 dpi, * p = 0.0209
120 dpi: mon. vs. PFF, * p = 0.0125
4f
Two-way ANOVA
Treatment x Time
F(2,29) = 0.9799; P = 0.3874
Time factor
F(2,29) = 4.684; P = 0.0173
Treatment factor
F(1,29) = 1.089; P = 0.3053
PFF: 0 dpi vs. 120 dpi, * p = 0.0343
4g
For 0, 60, 120 dpi:
α-Syn mon. = 9, 6, 4
α-Syn PFF. = 10, 7, 6
Two-way ANOVA
Treatment x Time
F(2,36) = 8.681; P = 0.0008
Time factor
F(2,36) = 4.437; P = 0.0190
Treatment factor
F(1,36) = 31.53; P < 0.0001
PFF: 0 dpi vs. 60 dpi, * p = 0.0110
PFF: 0 dpi vs. 120 dpi, *** p = 0.0002
60dpi: mon. vs. PFF, ** p = 0.0044
120 dpi: mon. vs. PFF, *** p = 0.0004
4h
Two-way ANOVA
Treatment x Time
F(2,36) = 2.229; P = 0.1223
Time factor
F(2,36) = 2.760; P = 0.0767
Treatment factor
F(1,36) = 3.591; P = 0.0661
PFF: 0 dpi vs. 120 dpi, * p = 0.0442
4i
Two-way ANOVA
Treatment x Time
F(2,36) = 1.724; P = 0.1927
Time factor
F(2,36) = 10.32; P = 0.0003
Treatment factor
F(1,36) = 1.917; P = 0.1747
PFF: 0 dpi vs. 120 dpi, *** p = 0.0006
4l
All conditions = 4
Two-way ANOVA
Treatment x Time
F(2,18) = 4.220; P = 0.0314
Treatment factor
F(1,18) = 15.22; P = 0.0005
Time factor
F(2,18) = 12.10; P = 0.0010
PFF: 0 dpi vs 120 dpi, *** p = 0.0005
120 dpi: PFF vs. mon., * p = 0.0102
4m
All conditions = 4
Two-way ANOVA
Treatment x Time
F(2,18) = 0.7729; P = 0.4764
Treatment factor
F(1,18) = 10.55; P = 0.0045
Time factor
F(2,18) = 5.665; P = 0.0124
PFF: 0 dpi vs. 120 dpi, p = 0.0771
Mon 0 dpi vs PFF 120 dpi, ** p = 0.0049
4n
For 0, 60, 120 dpi:
WT = 4, 4, 4
Aged PFF = 5, 5, 6
For 60, 120 dpi:
Aged mon. = 4, 4
ASO young = 4
ASO 12 m.o. = 6
One-way ANOVA F(9,36) = 6.176; P < 0.0001
Aged PFF: 0 dpi vs. 120 dpi, * p = 0.0487
ASO young vs. ASO 12 m.o., ** p = 0.0017
Extended Data Figures
Figure
Number of subjects
Test
Summary
Key comparisons
e1a
WT = 42
ASO = 20
Aged = 19
One-way ANOVA F(2,78) = 7.080; P = 0.0015
WT vs. ASO, ** p = 0.022
ASO vs. Aged, ** p = 0.0075
e1b
One-way ANOVA F(2,78) = 25.12; P < 0.0001
WT vs. ASO, **** p < 0.0001
WT vs. Aged, *** p = 0.0009
ASO vs. Aged, **** p < 0.0001
e1c
One-way ANOVA F(2,78) = 27.16; P < 0.0001
WT vs. ASO, **** p < 0.0001
ASO vs. Aged, ****p < 0.0001
e1d
WT = 13
ASO = 9
Aged = 12
One-way ANOVA F(2,31) = 14.69; P < 0.0001
WT vs. ASO, **** p < 0.0001
ASO vs. Aged, ** p = 0.0038
e1e
WT = 42
ASO = 20
Aged = 19
One-way ANOVA F(2,78) = 116.9; P < 0.0001
WT vs. Aged, **** p < 0.0001
ASO vs. Aged, **** p < 0.0001
e1f
WT = 3
ASO = 4
Aged = 4
One-way ANOVA F (2, 8) = 2.541; P = 0.1399
e1g
See Fig. 1e
e1i
All conditions = 4
One-way ANOVA F (7, 24) = 4.712; P = 0.0019
WT vs. ASO, * p = 0.0304
WT vs. PFF 60 dpi, * p = 0.0327
60 dpi: PFF vs. mon., * p = 0.0480
e2d
WT = 4
All PFF = 5
Mon. 7 dpi = 4
Mon. 60 dpi = 5
One-way ANOVA
Neurons per crypt
F (8, 45) = 0.07617; P = 0.9997
EGCs per crypt
F (8, 39) = 2.501; P = 0.0269
EGCs, WT vs. PFF 60 dpi, * p = 0.0329
EGCs, WT vs. PFF 120 dpi, *p = 0.0232
e2f
α-Syn mon. = 4
α-Syn PFF. = 5
Student’s t-test,
one-tailed
F(4,3) = 58.59; P = 0.0071
Mon. vs. PFF, * p = 0.0071
e3b
WT = 8
PFF 7 dpi = 5
PFF 21 dpi = 5
PFF 60 dpi = 5
PFF 120 dpi = 5
Mon. 7 dpi = 4
Mon. 60 dpi = 4
ASO = 8
Aged = 5
One-way ANOVA F (8, 40) = 5.132; P = 0.0002
WT vs. ASO, ** p = 0.0092
e3d
All conditions = 3
except BSA, 50 = 4
One-way ANOVA F (4, 11) = 7.188; P = 0.0042
BSA vs. PFF, 50, ** p = 0.0080
BSA vs. PFF, 100, *** p = 0.0009
Mon., 50 vs. PFF, 50, * p = 0.0178
Mon., 100 vs. PFF, 100, ** p = 0.0078
e4a
See Fig. 2k
e4b
e4c
e4d
e4e
All conditions = 4
Two-way ANOVA
Treatment x Genotype
F(2,18) = 0.8512; P = 0.4434
Time factor
F(2,18) = 0.4326; P = 0.6554
Genotype factor
F(1,18) = 12.10; P = 0.0027
e4f
Two-way ANOVA
Treatment x Genotype
F(2,18) = 0.08766; P = 0.9165
Time factor
F(2,18) = 0.8450; P = 0.4459
Genotype factor
F(1,18) = 6.227; P = 0.0225
e4g
Two-way ANOVA
Treatment x Genotype
F(2,18) = 0.5592; P = 0.5813
Time factor
F(2,18) = 0.5461; P = 0.5885
Genotype factor
F(1,18) = 9.940; P = 0.0055
e4h
Two-way ANOVA
Treatment x Genotype
F(2,18) = 0.1810; P = 0.8359
Time factor
F(2,18) = 0.4914; P = 0.6198
Genotype factor
F(1,18) = 41.38; P < 0.0001
0 dpi: WT vs. ASO, ** p = 0.0085
60 dpi: WT vs. ASO, * p = 0.0293
e5a
See Fig. 3i
e5b
See Fig 3j
e6c
WT = 5
For 7, 60, 120 dpi:
α-Syn PFF = 5, 5, 4
For 7, 60 dpi:
α-Syn mon. = 4, 3
ASO = 5
Aged = 5
One-way ANOVA F(7,28) = 5.007; P = 0.0009
WT vs. ASO, ** p = 0.0011
ASO vs. mon. 7 dpi, ** p = 0.0027
ASO vs. mon. 60 dpi, * p = 0.0176
ASO vs. Aged, * p = 0.0323
e7e
WT = 3
α-Syn PFF, 60 dpi = 3
ASO = 3
One-way ANOVA F(2, 6) = 47.90; P = 0.0002
WT vs. ASO, *** p = 0.0003
PFF 60 dpi vs. ASO, *** p = 0.0007
e7f
One-way ANOVA F(2, 6) = 32.76; P = 0.0006
WT vs. ASO, ** p = 0.0011
PFF 60 dpi vs. ASO, ** p = 0.0014
e8a
For 0, 7, 21, 60, 90, 120 dpi:
α-Syn PFF = 16,14,14,11,9,8
α-Syn mon. = 9,9,9,8,8,8
BSA = 17,16,11,9,7,7
Two-way ANOVA
Treatment x time
F(10,172) = 2.399; P = 0.0108
Time factor
F(5, 172) = 2.388; P = 0.0400
Treatment factor
F(2, 172) = 18.24; P < 0.0001
PFF: 0 dpi vs. 60 dpi, ** p = 0.0012
90 dpi: PFF vs. mon., * p = 0.0265
e8b
Two-way ANOVA
Treatment x time
F(10,172) = 1.215; P = 0.2844
Time factor
F(5,172) = 8.351; P < 0.0001
Treatment factor
F(2,172) = 8.086; P = 0.0004
PFF: 0 dpi vs. 60 dpi, * p = 0.0306
PFF: 0 dpi vs. 120 dpi, * p = 0.0285
e8c
Two-way ANOVA
Treatment x time
F(10,172) = 1.215; P = 0.2844
Time factor
F(5,172) = 8.351; P < 0.0001
Treatment factor
F(2,172) = 8.086; P = 0.0004
PFF: 0 dpi vs. 90 dpi, ** p = 0.0014
PFF: 0 dpi vs. 120 dpi, * p = 0.0285
60 dpi: PFF vs. mon., * p = 0.0193
90 dpi: PFF vs. mon., * p = 0.0342
e8d
For 0, 7, 21, 60, 90, 120 dpi:
α-Syn PFF = 8,8,8,12,8,8
α-Syn mon. = 9,9,9,8,8,8
BSA = 17,16,11,9,7,7
Two-way ANOVA
Treatment x time
F(10,152) = 0.1109; P = 0.9997
Time factor
F(5,152) = 10.52; P < 0.0001
Treatment factor
F(2,152) = 7.809; P = 0.0006
e8e
For 0, 7, 21, 60, 90, 120 dpi:
α-Syn PFF = 8,8,10,7,6,6
α-Syn mon. = 9,9,9,8,8,8
BSA = 17,16,11,9,7,7
Two-way ANOVA
Treatment x time
F(10,145) = 0.5406; P = 0.8589
Time factor
F(5,145) = 0.8005; P = 0.5510
Treatment factor
F(2,145) = 5.321; P = 0.0059
e8f
Two-way ANOVA
Treatment x time
F(10,145) = 1.352; P = 0.2087
Time factor
F(5,145) = 1.628; P = 0.1562
Treatment factor
F(2,145) = 17.66; P < 0.0001
60 dpi: PFF vs. BSA, ** p = 0.0023
e8g
Aged baseline: 10
Aged monomer, 60 dpi: 4
Aged PFF, 60 dpi: 6
One-way ANOVA
F(2,17) = 0.07091
e9d
All conditions = 4
One-way ANOVA F(2,9) = 0.1824; P = 0.8362