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SM Patogenesi e impatto dei nuovi trattamenti sul sistema immunitario

Luca Battistini

Unità di Neuroimmunologia

Luca Battistini M.D., Ph.D.

“Sclerosi Multipla: patogenesi e impatto dei

nuovi trattamenti sul sistema immunitario”

Luca BattistiniUnità di Neuroimmunologia

http://www.hsantalucia.it/laboratorio-neuroimmunologia

Cos’è la Neuroimmunologia


Sclerosi Multipla

MS patients have autoantibodies and/or self reactive T cells which are responsible for demyelination


Nature Reviews Neuroscience 3, 705-714. 2002

I protagonisti della SM


Autoimmunity

• Dal 7% al 10% degli adulti sonomalati nel nostro paese.

• Due terzi degli adulti malati sonodonne.

• Esistono più di 150 malattieautoimmuni/disimmuni e sonotriplicate negli ultimi 30 anni


Goodpasture’s syndromeGranulomatosis with Polyangiitis (GPA) see Wegener'sGraves' diseaseGuillain-Barre syndromeHashimoto's encephalitisHashimoto’s thyroiditisHemolytic anemiaHenoch-Schonlein purpuraHerpes gestationisHypogammaglobulinemiaIdiopathic thrombocytopenic purpura (ITP)IgA nephropathyIgG4-related sclerosing diseaseImmunoregulatory lipoproteinsInclusion body myositisInsulin-dependent diabetes (type1)Interstitial cystitisJuvenile arthritisJuvenile diabetesKawasaki syndromeLambert-Eaton syndromeLeukocytoclastic vasculitisLichen planusLichen sclerosusLigneous conjunctivitisLinear IgA disease (LAD)Lupus (SLE)Lyme disease, chronicMeniere’s diseaseMicroscopic polyangiitisMixed connective tissue disease (MCTD)Mooren’s ulcerMucha-Habermann diseaseMultiple sclerosisMyasthenia gravisMyositis

9

Acute Disseminated Encephalomyelitis (ADEM)Acute necrotizing hemorrhagic leukoencephalitisAddison's diseaseAgammaglobulinemiaAlopecia areataAmyloidosisAnkylosing spondylitisAnti-GBM/Anti-TBM nephritisAntiphospholipid syndrome (APS)Autoimmune angioedemaAutoimmune aplastic anemiaAutoimmune dysautonomiaAutoimmune hepatitisAutoimmune hyperlipidemiaAutoimmune immunodeficiencyAutoimmune inner ear disease (AIED)Autoimmune myocarditisAutoimmune pancreatitisAutoimmune retinopathyAutoimmune thrombocytopenic purpura (ATP)Autoimmune thyroid diseaseAutoimmune urticariaAxonal & neuronal neuropathiesBalo diseaseBehcet’s diseaseBullous pemphigoidCardiomyopathyCastleman diseaseCeliac diseaseChagas diseaseChronic fatigue syndrome**Chronic inflammatory demyelinating polyneuropathy (CIDP)Chronic recurrent multifocal ostomyelitis (CRMO)

Celiac diseaseChagas diseaseChronic fatigue syndrome**Chronic inflammatory demyelinating polyneuropathy (CIDP)Chronic recurrent multifocal ostomyelitis (CRMO)Churg-Strauss syndromeCicatricial pemphigoid/benign mucosal pemphigoidCrohn’s diseaseCogans syndromeCold agglutinin diseaseCongenital heart blockCoxsackie myocarditisCREST diseaseEssential mixed cryoglobulinemiaDemyelinating neuropathiesDermatitis herpetiformisDermatomyositisDevic's disease (neuromyelitis optica)Discoid lupusDressler’s syndromeEndometriosisEosinophilic fasciitisErythema nodosumExperimental allergic encephalomyelitisEvans syndromeFibromyalgia** Fibrosing alveolitisGiant cell arteritis (temporal arteritis)GlomerulonephritisGoodpasture’s syndromeGranulomatosis with Polyangiitis (GPA) see Wegener'sGraves' diseaseGuillain-Barre syndromeHashimoto's encephalitis

Meniere’s diseaseMicroscopic polyangiitisMixed connective tissue disease (MCTD)Mooren’s ulcerMucha-Habermann diseaseMultiple sclerosisMyasthenia gravisMyositisNarcolepsyNeuromyelitis optica (Devic's)NeutropeniaOcular cicatricial pemphigoidOptic neuritisPalindromic rheumatismPANDAS (Pediatric Autoimmune Neuropsychiatric Disorders Associated with Streptococcus)Paraneoplastic cerebellar degenerationParoxysmal nocturnal hemoglobinuria (PNH)Parry Romberg syndromeParsonnage-Turner syndromePars planitis (peripheral uveitis)PemphigusPeripheral neuropathyPerivenous encephalomyelitisPernicious anemiaPOEMS syndromePolyarteritis nodosaType I, II, & III autoimmune polyglandular syndromesPolymyalgia rheumaticaPolymyositisPostmyocardial infarction syndromePostpericardiotomy syndromeProgesterone dermatitisPrimary biliary cirrhosis

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Gray matter lesions

Immune system

Environmentalfactors

Genetic factors

Epigeneticfactors White matter lesions

Causes of Autoimmunity


14


1. CNS-extrinsic model

2.CNS-intrinsic model

(Adapted from Immunopathology of multiple sclerosis, Dendrou et al., Nat Rev Immunol 2015)

Autoreactive immune cells attack the CNS and cause neurodegeneration

Neurodegeration in the CNS causes infiltration of autoreactive immune cells

Periphery vs. CNS


1st ESCAPE of autoreactive immune cells from the thymus

Immunopathology of multiple sclerosis

(Adapted from Immunopathology of multiple sclerosis Calliope et al., Nat Rev Immunol 2015)

2nd ESCAPE of autoreactive

immune cells in the periphery

ACTIVATION of autoreactive immune cells

in the periphery

MIGRATION of autoreactive immune cells

in the CNS

Dendritic

cells


Regulation Inflammation

Immunological equilibrium


Immunological disequilibrium


Regulation Inflammation

Immunological equilibrium

https://www.daisychung.com/immunology-cartoon-project-


Nature Reviews Immunology 2005 5, 760 - 771

What is CD39?


CD39

ATP ADP AMP AMP

Adenosine

CD73

CD73


Megakaryoblast

Megakaryocyte

Thrombocytes

Polychromatic erythrocyte

(Reticulocyte)

Erythrocyte

Basophil Neutrophil Eosinophil Monocyte

Myeloid dendritic cellMacrophage

Multipotential hematopoietic stem cell(Hemocytoblast)

Common myeloid

progenitor

Common lymphoid

progenitor

Small lymphocyte

B cellNKT cell

Prolymphocyte

Lymphoid dendritic cell

CD4+ CD8+

T cell

Proerythroblast(Pronormblast)

Myeloblast

Plasma cell

Meet the players


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dic. ’18Courtesy of Mario Roederer 29

Diversity of Leukocytes

T CellsB CellsNK CellspDCMonocyte

s

mDC

DPCD4 CD8 DNVd1 Vd2NKT MAIT


dic. ’18 30


s

mDC

Th1 Th2 Th9 Th17 Th22 TREG TFH

CD4 T Cell “Lineages”

Courtesy of Mario Roederer



dic. ’18 31


s

mDC

SCM

Central

Transitional

Effector

Terminal

Naive




dic. ’18 32

T CellsB CellsNK CellspDCMonocytes mDC

Lineages

Dif

fere

nti

ati

on

/Eff

ecto

r F

un

cti

on



M1/M2 Macrophages

Macrophage

Myeloid DC

Th1

(IFN-γ)

Th17

(IL-17) Th2

(IL-4, IL-5,

IL-10, IL-13)

T reg

(TGF-β, IL-10)

Th1 responses Pathogens killing

Tumor resistance

Th2 responses Parasites killing

Th2 activation Immunoregulation

Immunoregulation Tissue remodeling

M2c

IL-10

M2b

IL-1β + LPS

M2a

IL-4

M1

IFN-γ + LPS

Inflammation Intracellular pathogens

Tumor resistance

Extracellular bacteria Fungi

Autoimmunity

Immunoregulation Extracellular parasites

Allergy and asthma

Tumor promotion

Immune tolerance Downregulation of

immune responses

TGF-β

IL-4

IL-12 TGF-β IL-6

IL-23

Plasmacytoid DC

Innate

compartment

Adaptive

compartment

monocyte


Dendritic cells

Macrophage

Myeloid DC

Th1

(IFN-γ)

Th17

(IL-17) Th2

(IL-4, IL-5,

IL-10, IL-13)

T reg

(TGF-β, IL-10)

Th1 responses Pathogens killing

Tumor resistance

Th2 responses Parasites killing

Th2 activation Immunoregulation

Immunoregulation Tissue remodeling

M2c

IL-10

M2b

IL-1β + LPS

M2a

IL-4

M1

IFN-γ + LPS

Inflammation Intracellular pathogens

Tumor resistance

Extracellular bacteria Fungi

Autoimmunity

Immunoregulation Extracellular parasites

Allergy and asthma

Tumor promotion

Immune tolerance Downregulation of

immune responses

TGF-β

IL-4

IL-12 TGF-β IL-6

IL-23

Plasmacytoid DC

Innate

compartment

Adaptive

compartment

monocyte


Dendritic Cells

In peripheral blood

CD123-CD11c+ myeloid dendritic cells

(mDC)

CD123+CD11c- plasmacytoid dendritic

cells (pDC)

Th-2

Th-17

B cells

Th-1Immunity

Tolerance

In tissues

IL-12

IL-1, TNF-a, IL-6

IL-1, TNF-a

IL-6

IL-4, IL-5


Markers for monitoring disease 1

36

cytometric analysis on freshly isolated PBMCs from healthy donors

and MS patients in different phasesof disease activity. Since CTLA4

and GITR staining display lower discriminating sensitivity, we used

CD39, FoxP3, and CD25 aspreviously described [38]. Indeed, stable

MS patientsshowed a significant reduction of Treg cellsascompared

to healthy donors (Fig. 3A–B; Fig. 4A–C). Samples collected from

patients during an acute attack, as suggested by mRNA levels,

displayed restored levelsof Treg cells, comparable to that observed in

healthy donors (Fig. 3B–C; Fig. 4A–C). We confirmed that

CD4+CD25highCD39+ T cells from MS patients in the acute phase

of the disease are indeed suppressive, as shown in Fig. 4 D–F, thussuggesting that the T regulatory compartment is not functionally

compromised in patients affected by MS.

Longitudinally followed MS patients display increase

Treg markers if experiencing a relapseWe then analyzed by RT-PCR, RNA samples from 15

untreated MS patients that had been followed longitudinally for

14 monthswith bi-monthly sampling, constituting theplacebo armof a clinical trial. While seven patients remained relapse-free

Figure 2. Treg markers are up-regulated in RR-MS pat ients experiencing clinical relapses. A–E. Clinically relapsing RR-MS patientsdisplayed increased PBMC mRNA levels for CD25 (A), CTLA-4 (B), GITR(C), CD39 (D), and foxp3 (E). We also found significantly lower levels of Tregmarkers mRNA in stable RR-MS patients as compared to healthy controls (HC). Values are expressed as arbitrary units (AU). P values are indicated(Mann-Whitney).doi:10.1371/journal.pone.0021386.g002

T Regulatory Cells in MS

PLoS ONE | www.plosone.org 4 June 2011 | Volume 6 | Issue 6 | e21386

Borsellino G. et al.Blood. 2007 Aug 15;110(4):1225-32.

Dalla Libera D.PLoS One. 2011;6(6):e21386..

100

101

102

103

104

100

101

102

103

104

100

101

102

103

104

3.65

18.8

16.7

3.65

18.8

16.7

CD4

CD

25

CD39


37


Annibali V. et al.Brain. 2011 Feb;134(Pt 2):542-54.

A CB

% MAIT/CD8


38


Chiurchiù V. et al.Annals of Neurology. 2013 May;73(5):626-36.

0 10K 20K 30K

FS L in: FS

0

10K

20K

30K

SS

Lin

: S

S

0 10 1

10 2

10 3

CD14

0

10 1

10 2

10 3

HL

A-D

R

0 10 1

10 2

10 3

CD3-CD56-CD19

0

10 1

10 2

10 3

HL

A-D

R

Distinct modulation of human myeloid and plasmacytoid dendritic cells by anandamide

in M ultiple Sclerosis

Valer io Chiurchiù 1,2, Mar ia Teresa Cencioni 1, Elisa Bisicchia 1,2, Marco de Bardi 1, Diego Centonze3, Mauro Maccarrone 1,2 and Luca Battistini1

1European Center for Brain Research/Fondazione Santa Lucia, Rome, Italy; 2Department of Biomedical Sciences, University of Teramo, Teramo, Italy;

3Department of Neurosciences, University of Rome “ Tor Vergata” , Rome, Italy.

• The immunopathogenesis of multiple sclerosis (MS) has always been thought to be driven by chronically activated and autoreactive Th-1 and Th-17 cells. Recently, also dendritic

cells (DC) have been thought to significantly contribute to antigenic spread and to maturation of adaptive immunity, and have been linked with disease progression and

exacerbation.

• To date, only a few studies have directly addressed the role of DC in MS patients, showing elevated numbers of peripheral blood DC with altered phenotype, and a dysfunctional

interaction with T cells. Yet, the exact phenotype and role of pDC and mDC during MS is not well understood

• The endocannabinoid system (ECS) has emerged as a potential target for MS management. Endocannabinoids have been shown to be neuroprotective and immunosuppressive.

MS patients bear increased levels of the main endocannabinoid anandamide (AEA) in CSF, lymphocytes and plasma (Centonze et al., 2007) and it inhibits human Th-1 and

Th-17 immune responses (Cencioni & Chiurchiù et al, 2010).

H S M S

TNF-α 54.2 ± 2.8 58.3 ± 3.2

IL-12 9.4 ± 0.8

19.3 ± 1.1 *

IL-6 13.9 ± 1.6

24.5 ± 1.9 *

H S M S

TNF-α

!52.1!± 2.2 56.4!±2.8

IFN-α

41.4!± 1.5 27.6!± 0.9 *

0 10 1

10 2

10 3

CD123

0

10 1

10 2

10 3

CD

11

c

mDC$

pDC$

H S M S

mDC

pDC

Differential cytokine production from healthy and MS dendritic cell subsets

R848-induced cytokine modulation by AEA in mDC and pDC Differential expression of ECS on mDC and pDC

In MS, mDC have

high CB2R (A) and

low FAAH (C)

mRNA levels,

whereas pDC show

unvaried CB2R (B)

and high FAAH (D)

mRNA levels.

CB2R

FAAH

CB2R protein was equally

expressed in healthy and

pathogenic pDC, whereas

FAAH was significantly

expressed in pathogenic

pDC

CONCLUSI ONS

In MS mDC produce higher levels of IL-12 and IL-6, whereas pDC account for

lower levels of IFN-α. Only pathogenic pDC lack responsiveness to cytokine

inhibition induced by AEA. Consistently, this specific cell subset expresses higher

levels of the anandamide hydrolase FAAH. These findings disclose a distinct

immunomodulation by AEA of mDC and pDC in MS patients, which may reflect

an alteration of the expression of FAAH, thus forming the basis for the rational

design of new endocannabinoid-based immunotherapeutics targeting a specific cell

subset.

AEA inhibits IL-12 and IL-6 production in both healthy and pathogenic mDC in

A CB2R-dependent manner

Pathogenic pDC are unresponsive to AEA-induced and CB2R-dependent IFN-α

modulation


Immuno-balance vs immunopathogenesisin multiple sclerosis

Th9 cells

B cells

monocytes

Th17 cells

Th1 cells

CD8 Tcells

Treg cells

Protective immune response

Pathogenic immune response


s.c.IFNb1b

1995 2000 2005 2010 2015 2017

i.m.IFNb1a

s.c.IFNb1a

GA 20

MITOX

ASCT

natalizumab fingolimod

teriflunomide

DMF

Peg IFNb1a

alemtuzumab

daclizumab

ocrelizumab

GA 40

cladribine

Terapie Attuali per laSclerosi Multipla

CYCLO

AZA


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17

Fig.%4:%Popolazioni%cellulari%implicate%nell’%immunopatogenesi%della%SM

3.2 I l ruolo delle cellule B nella SM

Diverse evidenze sperimentali suggeriscono che anomalie delle cellule B giochino un

ruolo chiave nella fisiopatologia della sclerosi multipla. Queste anomalie includono:

- elevati livelli di immunoglobuline prodotte all’interno del SNC (produzione

intratecale);

- presenza di bande oligoclonali IgG;

- accumulo di anticorpi e di complemento attivato legato alla mielina

danneggiata.

Alcune cellule B autoreattive riconoscono antigeni self. Nelle persone sane i processi

di sviluppo delle cellule comprendono dei checkpoint che limitano la produzione di

cellule B autoreattive. Nella SM, alcune cellule B autoreattive sono in grado di


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Loss of memory cells

IMMUNOSENESCENZA ED ESAURIMENTO

Schurich and Henson; Frontiers in Immunology

Loss of effector functions; loss of proliferative capacityretetention of cytotoxic activity, TNFα and IFNγ production


dic. ’18

Risposte ImmunitarieAutoimmunità

Allergia/Disimmunità

ImmunoregolazioneInfezioni croniche

Tumori

44Luca Battistini M.D., Ph.D.

dic. ’18 45

Mario Picozza Marco De Bardi Eleonora Piras Gloria DonninelliMarta Corsetti Francesca Gargano Elisabetta Volpe Roberta PlacidoGiovanna Borsellino Daniela F. Angelini Alessia Capone Manolo SambucciGisella Guerrera Francesca Candiloro Arianna Romoli Fortunata Carbone

Neuroimmunology Unit

Translational Cytomics Lab


Progetto realizzato con

il contributo non

condizionante di:

Fine


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