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CRAC Ca 2+ channels are essential for CD4 + T cell function in inflammatory bowel disease. Lymphocyte function is controlled by many ion channels. Lymphocyte function is controlled by many ion channels. Stefan Feske, MD , Department of Pathology New York University School of Medicine.

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CRAC Ca2+ channels are essential for CD4+ T cell function in inflammatory bowel disease

Lymphocyte function is controlled by many ion channels

Lymphocyte function is controlled by many ion channels

Stefan Feske, MD , Department of Pathology

New York University School of Medicine

CRAC (ORAI1, STIM1)

P2X

TRP?

Cav?

Kv1.3

KCa3.1

TRPM7

MagT1

Ca2+

K+

Ca2+

TRPM4

Na+

Mg2+

(Ca2+)

T cell

ZIP, ZnT

Zn2+

Cl–

Disclosure

CalciMedica: co-founder and consultant


S. typhi.

SFB

B. fragilis

DC

MF

Naïve

CD4

Naïve

CD4

Naïve

CD4

IL-23

TGFb

IL-12

IL-23R*

Treg

Foxp3

Th1

Th17

IL-21*

IL-10

TGFb

IL-17A

IL-17F

IL-22

CCR6*

IFN-g*


Store-Operated Ca2+ Entry (SOCE)

S. typhi.

SFB

B. fragilis

Antigen

DC

MF

Naïve

CD4

Ca2+

Naïve

CD4

Naïve

CD4

IL-23

TGFb

IL-12

IL-23R

Treg

Foxp3

Th1

Th17

IL-21

IL-10

TGFb

NFAT

ERK

NFkB

IL-17A

IL-17F

IL-22

CCR6

IFN-g


CRAC channel pore protein ORAI1 is required in CD4+ T cells

to cause IBD in mice

CD45RB T cell transfer model of colitis

Weight loss

Isolate naïve CD4+ T cells

(CD45RBhi CD25neg)

+/+

+/+

+/+

+/+

KI/KI

KI/KI

IBD

WT or CRAC-deficient mice

Rag2-/- mice

(no T, B or Treg cells)

Weeks after T cell transfer

Orai1KI/KI

WT

WT

Orai1KI/KI


Ca2+ influx levels in CD4+ T cells determine severity of colitis

CD4+ T cells

WT

[Ca2+]i

Orai1KI

Stim1fl/fl

Weight loss after CD4+ T cell transfer

min

Stim2fl/fl

Orai1fl/fl

WT

[Ca2+]i

WT

Stim1fl/fl

Cd4Cre

Cd4Cre

min

ER

Weeks

Nucleus

WT

Stim1fl/fl

Cd4Cre

[Ca2+]i

WT

Stim2fl/fl

Cd4Cre

min


Ca2+ influx levels in CD4+ T cells determine severity of colitis

WT

[Ca2+]i

Orai1KI

Stim1fl/fl

Weight loss after CD4+ T cell transfer

min

Stim2fl/fl

Orai1fl/fl

WT

[Ca2+]i

WT

Stim1fl/fl

Cd4Cre

Cd4Cre

min

Ca2+

WT

ER

Weeks

Stim1fl/fl

Nucleus

Stim2fl/fl

Colitogenic

Threshhold

[Ca2+]i

WT

Orai1fl/fl

Stim2fl/fl

Cd4Cre

time

min


Ca2+ signaling through CRAC channels is not required for

Proliferation, survival or gut homing of CD4+ T cells

MFI

Lamina

propria

MFI

  • Normal numbers of CD4+ T cells (including Foxp3+Treg cells)

  • Normal expression of T cell homing molecules

  • Normal apoptosis and proliferation rates

    • in lamina propria (colon) and mesenteric lymph nodes


SOCE levels determine IFNgand IL-17 production

in lamina propria CD4+ T cells

Lamina

propria

MFI

Cytokine production

by CD4+ T cells in lamina propria


SOCE is required for Th17 cell function & differentiation

IL-17A

IL-17F

IL-22

Th17

Naïve

CD4+ T cell

Th17

TGF-b

IL-6

RORgt

RORgt

In vitro differentiatedTh17 cells


SOCE is required for Th17 cell function & differentiation

IL-17A

IL-17F

IL-22

Th17

Naïve

CD4+ T cell

Th17

TGF-b

IL-6

RORgt

RORgt

CCR6

IL-23R

Terminal

differentiation

IL-21R

IL-21

IL-23

In vitro differentiatedTh17 cells

(from DC)

IL-21

IL-23R

CCR6

% CCR6+ CD4+

*

WT

IL-23R

Stim1fl/flCd4Cre

Actin

iso

WT

WT

WT

WT

Stim1fl/flCd4Cre

Stim1fl/fl Cd4Cre

Stim1fl/fl Cd4Cre

Stim1fl/flCd4Cre


CRAC channel inhibition as anti-inflammatory treatment in IBD ?

Inducible genetic

Deletion using

Stim1fl/fl ERT2Cre mice

ERT2-Cre

Tamoxifen

Cre

TAM

nucleus

Stim1fl/fl

Stim1-/-

– TAM

+ TAM


Deletion of STIM1 (SOCE) IBD ?in T cells after IBD development

ameliorates intestinal inflammation

Transfer of CD4+ CD45RB+ T cells from Stim1fl/fl ERT2Cre mice

Vehicle (+ STIM1)

Tamoxifen (– STIM1)

Treg

(Foxp3+)

% CD4+ CD25+ Foxp3+

+ STIM1

– STIM1


Summary IBD ?

CRAC channels (ORAI1, STIM1, STIM2)

in CD4+ T cells:

Essential for Ca2+ influx

Induction of IBD

Th1 and Th17 function:

production of proinflammatory cytokines (IL-17A, IL-17F, IL-22,

IFNg, TNFa) in the gut

Terminal Th17 differentiation

Severity of IBD correlates with

levels of Ca2+ influx and cytokine production

Induced genetic deletion of

STIM1 in T cells mimics CRAC

inhibitors and leads to rapid

recovery from IBD

S. typhi.

SFB

B. fragilis

DC

MF

Naïve

CD4

Naïve

CD4

Naïve

CD4

IL-23

TGFb

IL-12

IL-23R

Treg

Foxp3

Th1

Th17

Ca2+

IL-21

IL-10

IL-17A

IL-17F

IL-22

CCR6

Ca2+

IFN-g

Ca2+

Adapted from: Arrieta

Front Immunol (2012)


Carl IBD ?Weidinger

Patrick Shaw

Christie-Ann McCarl

Sarah Khalil

JianMa

NIH/NIAID