Interplay of subclinical fibrosisand

inflammation

D. SerónNephrology Department Hospital Vall d’HebronBarcelona

Inflammation with and without fibrosis

Inflammation and CHR

Innate immunity and inflammation

Surveillance biopsies

Inflammation with and without fibrosis

Inflammation and CHR

Innate immunity and inflammation

Surveillance biopsies

Inflammation

Inflammation and fibrosis in

surveillance Bx

Dimény E, Clin Transplantation 1995; 58(11): 1195Serón D, Kidney Int 1997; 51: 310Nankivell BJ et al, Transplantation 2004; 78:242Choi BS et al, Am J Transplant 2005; 5: 1354

Fibrosis

Graft survival

Graft survival

SCR and CAN in paired biopsiesn=598 Bx, (no SCR 462, SCRB 102, SCRA 34)

Nankivell BJ et al, Transplantation 2004; 78:242Ibernon et al et al, Kidney Int 2006; 76: 557

* p<0.05***p<0.001

Proper classification of inflammation and fibrosis

Normal (no inflammation no fibrosis)

Inflammation and fibrosis

Inflammation (no fibrosis)

Fibrosis (no inflammation)

SCR & IF/TA

Shishido et al, JASN 2003; 14: 1046Cosio FG et al, Am J Transplant 2005; 5: 2464, Moreso F et al Am J Transplant 2006; 6:747

IF/TA without SCR

IF/TA with SCR

Normal1 year protocol BxSCR + IF/TA

SCR + IF/TA

SCR + IF/TA

1 year protocol Bx

.25

.5

.75

1

0 50 100 150 200 months

Normal=186

SCR=74

IF/TA=110

IF/TA+SCR=65

< 6 month protocol Bx

IF/TA without SCR

IF/TA with SCR

Hig

h in

flam

matio

n

Low

infla

mm

atio

n

CD45 CD3 CD20 CD68 CD15

High (upper tertile) and low (two lower tertiles) inflammation

Inflammation with or whitout IF/TA

Moreso F et al. AJT 2007; 7: 2739

0

500

1000

1500

2000

2500

NORMAL SCRa IF/TA SCR+IF/TAa,c0

300

600

900

1200

1500

NORMAL SCRa IF/TA SCR+IF/TAa,c

0

200

400

600

800

NORMAL SCR IF/TAa SCR+IF/TAa,b,c 0

500

1000

1500

2000

2500

3000

3500

NORMAL SCRa IF/TA SCR+IF/TAa,c

CD

45 p

osit

ive c

ells /

mm

2 in

ters

titi

um

CD

20 p

osit

ive c

ells /

mm

2 in

ters

titi

um

CD

68 p

osit

ive c

ells /

mm

2 in

ters

titi

um

CD

3 p

osit

ive c

ells /

mm

2 in

ters

titi

um

Interstitial infiltrating cells

Moreso F et al. AJT 2007; 7: 2739

Renal allograft survival(univariate)

0

,2

,4

,6

,8

1

Survival (%)

0 20 40 60 80 100 120 140

months

T1T2

T3

Moreso F et al Am J Transplant 2006; 6: 747

P=0.001

Renal allograft survival(multivariate)

Variable Univariate MultivariateRR (95% CI) p RR (95% CI) p

Retransplant (yes) 2.5 (0.9-6.9) 0.06 1.2 (0.3-5.3) ns

PRA > 50% 4.6 (1.7-12.6) <0.01 4.5 (1.0-20.4) 0.05

CD20 (upper tertile) 3.3 (1.4-8.3) <0.01 3.0 (1.2-7.3) 0.01

Moreso F et al Am J Transplant 2006; 6: 747

B cells, acute rejection and outcome inindication biopsies

Sarwal et al. et al. NEJM 2003; 349: 125.Hippen BE et al. Am J Transplant 2005; 5: 2248Alausa M et al. Clin Transplant 2005; 10: 137Eileen WT et al. Transplantation 2006; 82: 1769Lehnhardt et al. Am J transplant 2006; 6: 847Zarkhin V et al. Kidney Int 2008; 74: 664Mourah MR et al. Pediatr Transplant 2009; 13

B cells associated with poor outcome in AR

B cells not associated with poor outcome in ARScheepstra C et al. Transplantation 2008; 86:772Bagnasco SM et al. Am J Transplant 2007; 7: 1968-1973

Immunophenotype in protocol biopsies from TAC vs CsA treated patients

n= 44TAC vs 22 CsA

P<0.01

P<0.01 P<0.05ns

Serón D et al, Transplantation 2007; 83:649

B cell survival in CHR after rituximabn=38 explanted grafts with CHR 2 after treatment with rituximab

Intragraft B cells

BAFF positivity

Thaunat et al. Transplantation 2008; 85:1648

SCR vs SCR+IF/TA and FoxP3

Bestard et al. JASN 2008; 19: 2010

SCR SCR +No IF/TA IF/TA p

N 19 18___________________________________________________%FoxP3 + cells 14.2 +/-16.3 8.2+/-8.1 0.035___________________________________________________

T regs in pts ACR and BL (indication (12 ACR, 12 BL) and surveillance (8ACR, 8 BL)

Taflin et al. Transplantation 2010; 89: 194

Surveillance Bx cytokine transcripts

Hueso M et al. Am J Pathol 2010; 176: 1696

P< 0.01

IL10

Inflammation+IFTA

B cellsTregs

Th2 responseAg presentation

Persistent inflammation?CHR?

Inflammation with and without fibrosis

Inflammation, IF/TA and CHR

Innate immunity and inflammation

Surveillance biopsies

Tx Inflammation CAN±

Inflammation

Survival

SCR and CHR

Tx Inflammation IF/TA±

Inflammation

Survival

SCR and CHR

CHR

SCR and Chronic humoral rejection1988-2006

Protocol Bx n = 517

CHR 44IF/TA nos 42Recurrence 11De novo GN 7Acute rejection 4Polyoma 1

Bx for cause: n = 109

Clinical characteristics at the time of biopsy

Variable CHR (44) IF/TA (42) p

Protocol biopsyTime (m) 4.5 ± 2.4 4.6 ± 3.3 nsSCr (mol/L) 149 ± 37 144 ± 44 nsProteinuria (g/d) 0.3 ± 0.2 0.3 ± 0.2 ns

Biopsy for causeTime (y) 6.4 ± 3.4 8.2 ± 4.4 0.037SCr (mol/L) 240 ± 141 204 ± 80 nsProteinuria (g/d) 2.3 ± 2.4 1.4 ± 1.7 0.061

Clinical characteristics of patientsVariable CHR IF/TA p-value

(n=44) (n=42)__________________________________________________Donor age (years) 40 ± 16 34 ± 15 nsDonor gender (% male) 65.9 78.6 nsPatient age (years) 43 ± 12 40 ± 12 nsPatient gender (male) 59.1 69.0 nsPRA (%) 7 ± 18 3 ± 10 nsVirus hepatitis C 18.2 9.5 nsRe-transplants (%) 15.9 2.3 0.058HLA DR mm 0.7 ± 0.5 0.7 ± 0.6 ns

Cold ischemia time (hours)23 ± 6 21 ± 6 nsImmunosuppression

CNI without MMF 24 27CNI with MMF 18 12CNI with mTOR-i 1 2CNI free 1 1 ns

Delayed graft function (%) 25 19 nsAcute rejection (%) 22.7 26.2 ns_________________________________________________________

(p=0.003)

Acute score

Acute Banff score in surveillance Biopsies

SCR, CHR and IF/TA

CHR IF/TA p__________________________________SCR (%) 52.3 28.6 0.025__________________________________

RR 95% CI p__________________________________SCR 2.52 1.1-6.3 0.047ReTx 6.7 0.8-58.8 ns__________________________________

n=34 n=17 n=17 n=18

% of cases with CHR and IF/TA in the biopsy for cause

Dg categoryin surveillance Bx

Is inflammation in SCR different from SCR + IFTA?

Are early Bx findings different in patients developing IFTA in comparison with patients developing CHR

Innate immune alterations and inflammation

Surveillance biopsies

Risk factors associated with early inflammation in surveillance biopsies

Immunosuppressive treatment

Acute rejection before surveillance Bx

Innate immunity & inflammation in the general population

TLR-2 expression in 6 m surveillance Bx is associated wit lower SCR257 surveillance Bx 6w, 3m, 6m and 108 indication Bx

De Groot K et al. Am J Nephrol 2008; 28: 583

MBL 96KDa protein made of 3 identical 32 KDa structures

Carbohydrate recognition domain

Collagen like domain

N-terminal cross linking region

N-acetylglucosamine D-mannoseN-acetyl mannosamineL-fructose

Bouwman LH et al. 2006; 67:247

Defense collagens

Bohlson SS et al. Mol Immunol 2007; 44:33

MBL in Renal TransplantsMar 2005 –Oct 2006, 125 RT, 111 with a functioning graft at 3 m

0

,5

1

1,5

2

2,5

3

3,5

4

Cel

l Mea

n

MBL-T1 MBL-T2 MBL-T3Cell

Interaction Bar Plot for CualquierMBLEffect: MBL-terciles

T 1

Low MBLN=42

T 2,3

High MBLN=83Log MBL

(ng/ml)

sTNFR2 before Txsoluble TNFR

p=0.05

Ibernon M et al. Transplantation 2009: 88: 272

MBL and infection (bacterial or fungal)March 2005-Oct 2006, 125 pts,

111 with a functioning graft at 3 m

Ibernon M et al. Transplantation 2009: 88: 272

MBL and NODAT March 2005-Oct 2006, 125 pts,

111 with a functioning graft at 3 m

Ibernon M et al. Transplantation 2009: 88: 272

P=0,0054

Low MBL and SCR(n=60)

Ibernon M et al. Kidney Int (in press)

(N=6)

(N=32)

Low MBL, subclinical rejection and Tx CAD

Arnt RF et al. Eur Heart J 2005; 26: 1660

C1q and MBL and rejection

Bohlson SS et al. Mol Immunol 2007; 44:33

C1q deficiency and acute rejectionHeart transplant in C1q deficient mice

Csencsits K et al. AJT 2008; 8: 1622

WT

C1q-/-

C1q deficiency and acute rejectionHeart transplant in C1q deficient mice

Csencsits K et al. AJT 2008; 8: 1622

T cell response is not enhanced in C1q-/- mice

More intense anti-donor Ab response

Tissue injury and repair

TxInjuryApotosisNecrosis

RepairNormalIFTA

Progression of injurySCR + IFTACHR

-

+

Inflammation(SCR)

Innnate Immunity