ApoE-/-hCD1Tg/HJ1Tg/

ApoE-/-hCD1Tg/HJ1Tg/

ApoE+/+

H&E(100x)

Supplementary Figure 1. No obvious pathology is observed in the colon of diseased hCD1Tg/HJ1Tg/ApoE-/-

mice. Colon samples were fixed in 10% formalin and laid out in Swiss rolls for paraffin embedding. Paraffin sections were cut and stained with H&E to examine the tissue architecture. Scale bars represent 100 µm.

15

10

5

0

2

1

0

Cervical Lymph Node Skin (dermis)

% C

D4+ F

oxp3

+

% C

D4+ F

oxp3

+

hCD1Tg/HJ1Tg/ApoE+/+

hCD1Tg/HJ1Tg/ApoE-/-

ApoE-/-

A B

Supplementary Figure 2. hCD1Tg/HJ1Tg/ApoE+/+ and hCD1Tg/HJ1Tg/ApoE-/- mice have comparable numbers of FoxP3+ regulatory T cells. Cells isolated from the (A) cervical lymph nodes and (B) dermis were stained intracellularly with Foxp3 and gated on CD45+TCRb+. Subsequently, the CD4 and Foxp3 positive Treg cell population was examined (n=3-5). ***P < 0.005; **P < 0.01; *P< 0.05; n.s. = not significant. Statistical analysis was performed using One-Way ANOVA followed by Bonferroni post-hoc test.

A B

0.00

6.37

0.15

19.2 5.35

2.49

0.53

6.32

0.00

3.33 4.43

5.92

0.00

0.84

0.00

4.62 3.61

4.43

CD1b

SSC

Gated on CD11b+CD11c+ cells

Gated on CD11b+CD11c- cells

Gated on CD11b-CD11c+ cells

CD

11b

CD11c

ApoE-/-hCD1Tg/HJ1Tg/

ApoE-/-hCD1Tg/HJ1Tg/

ApoE+/+

Dermis

0.890 0.82

CD1b

Gated on CD207+CD11c+ cells

SSC

Epidermis

8.183.38 7.79

CD

207

CD11c

ApoE-/-hCD1Tg/HJ1Tg/

ApoE-/-hCD1Tg/HJ1Tg/

ApoE+/+

Supplementary Figure 3. CD1b is expressed on a subset of dermal dendritic cells. (A) Dermal cells were isolated and stained with mAbs against CD45, CD11b, CD11c, and CD1b. CD1b expression on CD11b+CD11c-, CD11b+CD11c+ and CD11b-CD11c+ populations were analyzed by flow cytometry. (B) Epidermal cells were isolated and stained with mAbs against CD45, CD207, CD11c and CD1b. CD1b expression on CD207+CD11c+ population (Langerhans cells) was analyzed by flow cytometry by gating on CD45+ cells. (C) CD1b expression on hair follicle stem cells and keratinocytes was determined by gating on CD45- cells and then on integrin a6 and b1 double positive cells. Subsequently, CD34+ Sca-1- hair follicle stem cells or CD34- Sca-1+ epidermal keratinocytes were output for CD1b expression. Data shown are representative of at least 2 experiments for all figure panels.

0.54 0.630.25

00 0

Keratinocytes

Hair follicle stem cells

Epidermis

ApoE-/-hCD1Tg/HJ1Tg/

ApoE-/-hCD1Tg/HJ1Tg/

ApoE+/+

C

CD1b

SSC

Supplementary Figure 4. Fatty acids do not show preferential accumulation in the skin of hCD1Tg/HJ1Tg/ApoE-/- mice and reactivity of lipids to a second mouse CD1b-autoreactive hybridoma. Total lipid was extracted from skin and liver tissues of indicated mice, weighed and analyzed by gas chromatography. (A) Fatty acid species in the skin of diseased hCD1Tg/HJ1Tg/ApoE-/- and healthy hCD1Tg/HJ1Tg/ApoE+/+ mice (n=3) were quantified. (B) Ratio of fatty acids from diseased over healthy mice in the skin and liver. (C) Polar phospholipids (PL), cholesterol (Chol) and fatty acid mixtures (FA1: saturated and monounsaturated and FA2: polyunsaturated) were loaded on to CD1b protein and incubated with LN1-7 T cell hybridoma for 24 h. IL-2 in the supernatant was measured by ELISA. Data are representative of at least 2 experiments. **P < 0.01; *P< 0.05. Statistical analysis was performed using Student’s t-test.

0

2

4

32

34

with CD1b

LN1-7 reactivity to lipids

IL-2

Fol

d C

hang

e

C Lauric

Acid

Myristi

c Acid

Palmitic

Acid

Palmito

leic A

cid

Stearic

Acid

Oleic A

cid

Linoleic A

cid

Arachidic

Acid

a-Linolen

ic Acid

Eicose

noic Acid

Eicosa

dienoic

Acid

Behen

ic Acid

Arachidonic

Acid

Erucic Acid

Lignoceric

Acid

Nervonic

Acid

Docosa

hexae

noic Acid

0

1

2

3

4

5

Fold

Cha

nge

(Apo

E-/-/A

poE+/

+ )

Fatty acids fold diff

Skin

Liver

SkinLiverFold Change

nmol

/mg

dry

lipid

wei

ght

ApoE

-/-/A

poE+

A

B

Lauri

c

Myristi

c

Palmitic

Palmito

leic

Stearic

Oleic

Linole

ic

Arachid

ic

a-Lino

lenic

Eicose

noic

Eicosa

dieno

ic

Behen

ic

Arachid

onic

Erucic

Ligno

ceric

Nervon

ic

Docos

ahex

aeno

ic0

20406080

200400600800

1000nm

ol/m

g dr

y lip

id w

t.

Fatty acids skin

hCD1Tg+/HJ1Tg+/ApoE+/+

hCD1Tg+/HJ1Tg+/ApoE-/-

1000800600400200

806040200

5

4

3

2

1

0

*

**

**

*

*0.06 * *

hCD1Tg/HJ1Tg/ApoE+/+

hCD1Tg/HJ1Tg/ApoE-/-

Supplementary table 1: Oligonucleotide primers used in the studyName Sequence (5'-3')For genotypingHJ1a F TGACACCTGCTCAGTTCTTGTGCHJ1a R TAGCTTGTTCCCTGCACTTGG

CD1c F CAGAAACTGCAGAAACAGCACD1c R GATGGGTGAAAAGAGGTGAAA

ApoE WT TGTGACTTGGGAGCTCTGCAGCApoE Common GCCTAGCCGAGGGAGAGCCGApoE Mutant GCCGCCCCGACTGCATCT

For RT-PCRIL-18Ra F TCACCGATCACAAATTCATGTGG IL-18Ra R TGGTGGCTGTTTCATTCCTGT

IL-12Rb1 F TGCCGCTACTTCTCCTCAG IL-12Rb1 R ACTTCATGGTTCGGTTCCCAA

TLR9 F ATGGTTCTCCGTCGAAGGACTTLR9 R GAGGCTTCAGCTCACAGGG

S100a7 F GTA CTC AGG TCA TGG TTC TGS100a7 R GGT ATT CAA GCA AGG TAT CAC

S100a8 F GGA GTT CCT TGC GAT GGT GATS100a8 R TCC TTG TGG CTG TCT TTG TGA

b-actin F CTT CTT TGC AGC TCC TTC GTTb-actin R AGG AGT CCT TCT GAC CCA TTC

IL-17F F TGC TAC TGT TGA TGT TGG GACIL-17F R AAT GCC CTG GTT TTG GTT GAA

IL-17A F ACT TTC AGG GTC GAG AAG AIL-17A R TTC TGA ATC TGC CTC TGA AT

IL-22 F ATG AGT TTT TCC CTT ATG GGG ACIL-22 R GCT GGA AGT TGG ACA CCT CAA

GM-CSF F GGC CTT GGC AGC ATG TAG AGCGM-CSF R GGA GAA CTC GTT AGA GAC GAC TT

TNF-a F CCA CCA CGC TCT TCT GTC TTNF-a R GGC ACC ACT AGT TGG TTG T

IL-6 F CTT GGG ACT GAT GCT GGT GAC AIL-6 R GCC TCC GAC TTG TGA AGT GGT A

TGb1 F CTC CCG TGG CTT CTA GTG CTGb1 R GGCC TTA GTT TGG ACA GGA TCT G

IL-4 F AAC TCC ATG CTT GAA GAA GAA CTCIL-4 R CCA GGA AGT CTT TCA GTG ATG TG

IL-23 F CCA TTA GGA CTT GTG CTG TTC TIL-23 R CCA AGG GCT CGA GAC TTT ATT C

IFN-g F TCA AGT GGC ATA GAT GTG GAA GAAIFN-g R TGG CTC TGC AGG ATT TTC ATG

HJ1Vb F ATC TCT TCC CGG TGC TGA TTHJ1Vb R TCT GGT TCC TGA GCC AAA AT