DECREASED PERIPHERAL BLOOD BDCA1+ AND DECREASED PERIPHERAL BLOOD BDCA1+ AND BDCA3+ DENDRITIC CELLS IN RHEUMATOID ARTHRITIS BDCA3+ DENDRITIC CELLS IN RHEUMATOID ARTHRITIS

AND OSTEOARTHRITISAND OSTEOARTHRITISM VILLARROEL1, G ZALDIVAR1, MA SÁNCHEZ1, A HERNÁNDEZ1, JDD GARCÍA2, MJ LEÓN3, A SÁNCHEZ-ATRIO3, A

PÉREZ-GÓMEZ3, E CUENDE3, A LÓPEZ3, F ALBARRAN3, J MONSERRAT1, A PRIETO1, E REYES1, M ALVAREZ-MON1,3

(1) Laboratory of Immune System Diseases and Oncology. R&D CNB-CSIC associated unit. Department of Medicine, Alcalá de Henares, Spain. (2) Department of Internal Medicine. “Principe de Asturias” University Hospital, Alcalá de Henares, Spain.(3) Immune System Diseases and Oncology Service. “Principe de Asturias” University Hospital, Alcalá de Henares, Spain.

INTRODUCTION

Circulating Peripheral Blood Dendritic Cells (CPB-DCs) comprise different subsets that are involved into inflammatory response of patients suffering autoimmune and degenerative chronic diseases.

MATERIAL AND METHODS

Whole anticoagulated peripheral blood samples were obtained from RA patients (N=30), OA patients (N=21) and HC (N=7).

Blood Dendritic Cell Enumeration kit (Miltenyi Biotec – MACS ®) was used for phenotypical assay. We used four colours flow cytometer (FACSCalibur BD®) for data acquisition. CPB-DCs were stained with anti-BDCA1 (mDC1), anti-BDCA2 (pDC) and anti-BDCA3 (mDC2)

monoclonal antibodies in the CD19-CD14-EMA- population. Phenotypical and statistical analyses were performed with FlowJo (Tree Star®) and SPSS 11.0 (SPSS Inc.®) software respectively (Fig 1).

RESULTS

Significant decreases in the percentages of CD19-CD14-

EMA-BDCA1+ and CD19-CD14-EMA-BDCA3+ subsets were found in RA patients and OA patients compared to HC (RA: p<0.01 and p <0.04 respectively; OA: p<0.05 p<0.01 respectively) (Fig 2).

We found similar percentages of both mDC1 and mDC2 subsets between RA and OA patients. The percentage of

CD19-CD14-EMA-BDCA2+ DC subset did not differ in all three groups studied (Fig 3).

CONCLUSIONCONCLUSION

We found a significant decrease in mDC subset in We found a significant decrease in mDC subset in RA and OA patients compare to HC. RA and OA patients compare to HC.

This data also could be explained partially by the This data also could be explained partially by the chronic inflammatory response in both chronic inflammatory response in both pathologies. pathologies.

The decreased percentage of mDC subset found in The decreased percentage of mDC subset found in these patients may reflect either increased these patients may reflect either increased apoptosis of these cells or alternatively to apoptosis of these cells or alternatively to enhanced migration to sites of inflammation.enhanced migration to sites of inflammation.

AIM

The aim of this study is to compare the distribution of CPB-DC subsets in patients with rheumatoid arthritis (RA), osteoarthritis (OA) and healthy controls (HC).

Figure 1. Flow cytometric analysis of CPB-DCs.

The right contour plots showed the different CPB-DCs subsets, assessed in the CD19-CD14-EMA- population.

A.Sample stain with cocktail control.B.Sample stain with DBCA cocktail.Abreviations: FITC, fluorescein isothiocyanate; PE, phycoerithrin; PE-Cy5.5,

phycoerithrin-cyanin 5.5; APC, allophyco-cyanin. EMA. Ethidium Bromide Monoazide.

Figure 2. Distribution of mDC1 and mDC2 subsets in CPB-DCs.Percentage distribution. A. mDC1. anti BDCA1+

B. mDC2. anti BDCA3+

Numbers represent the significant values of U Mann Whitney test. Blue box differences between OA and HC, orange box differences between RA and HC.

RA. Rheumatoid arthritis. OA osteoarthritis. HC. Healthy controls.

Figure 3. Distribution of pDC subset in CPB-DCs

Percentage distribution.RA. Rheumatoid arthritis. OA

osteoarthritis. HC. Healthy controls

U.A.HU.A.H

HCOARA

pDC

,8

,6

,4

,2

0,0

-,2

HCOARA

mD

C 1

1,4

1,2

1,0

,8

,6

,4

,2

0,0

-,2

p<0.01

p<0.05

HCOARA

mD

C 1

1,4

1,2

1,0

,8

,6

,4

,2

0,0

-,2

p<0.01

p<0.05

HCOARA

mD

C 2

,2

,1

,1

0,0

-,1

p<0.04

p<0.01

HCOARA

mD

C 2

,2

,1

,1

0,0

-,1

p<0.04

p<0.01

A B

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

(A)

(B)

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

FL3H

: EM

A, C

D14

PE

-Cy5

.5, C

D19

PE

-Cy5

.5

(A)

(B)