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AB0473 Effects of CTLA4-IG (Abatacept) on Endothelial Cell Cultures After Different Times from Treatment
  1. M. Cutolo1,
  2. S. Soldano1,
  3. P. Contini2,
  4. A.C. Trombetta1,
  5. A. Sulli1,
  6. B. Seriolo1,
  7. M.A. Cimmino1,
  8. S. Paolino1,
  9. C. Pizzorni1,
  10. P. Montagna1,
  11. R. Brizzolara1
  1. 1Research Laboratory and Academic Division of Clinical Rheumatology, Department of Internal Medicine DIMI
  2. 2Laboratory of Clinical Immunology, Department of Internal Medicine DIMI, University of Genova, Genova, Italy

Abstract

Background Endothelial cell (EC) dysfunction is involved in the angiogenetic processes observed in rheumatoid arthritis (RA) synovitis [1]. CTLA4-Ig (abatacept) is employed as biological agent in RA treatment and interacts in vitro with the costimulatory molecule CD86 (B7.2) expressed by different cells involved in the process, including ECs [2-4].

Objectives To evaluate the expression of CD86, CD54 (ICAM1, intercellular adhesion molecule 1) and CD309 (VEGFR-2, vascular endothelial growth factor receptor 2) on cultured human microvascular endothelial cells (HMVECs), activated with γIFN, treated with abatacept (ABAT) and tested after different times from treatment.

Methods HMVECs (Lonza, Switzerland) were activated with γIFN (500 U/ml), treated with ABAT (10, 50, 100, 500 μg/ml) and evaluated after different times of incubation (24, 48, 72 hrs and 7 days). Evaluation of CD86, CD54, CD309 expression were performed at fluorescence-activated cell sorting analysis (FACS, FC500, Coulter, Hialeah, FL) immediately after the first activation with γIFN (control) and after each ABAT treatment (24, 48, 72 hrs and 7 days).

Results FACS analysis showed in activated HMVECs a decrease of the CD86+ cell percentage, after 24 and 48 hrs from ABAT (500 μg/ml) treatment (77% and 69% CD86+ cells, respectively) and after 48 hrs from ABAT (100 μg/ml) treatment (93% CD86+ cells), compared to the untreated controls (97% CD86+ cells). On the contrary, no reductions regarding the % of CD86+ cells were further detectable after 72 hrs and 7 days, compared to basal values. No changes were observed after other ABAT doses (10 and 50 μg/ml).

There was a reduction in the percentage of CD54+ cells at all time points after treatment with ABAT (48 hrs, 72 hrs and 7 days). Interestingly, after only 24 hours ABAT (100 and 500 μg/ml) induced a decrease of the absolute percentage of CD54+ cells (18% and 20% CD54+, respectively) compared to controls (42% CD54+ cells).

On the other hands, CD309+ cells showed a % decrease (71% and 41%, respectively) after 24 hrs from ABAT treatments (both 100 and 500 μg/ml) and after 48 hrs from ABAT (500 μg/ml) treatment (62% CD309+ cells), compared to the control (96% CD309+ cells). No changes for the % of CD309+ cells were observed after other ABAT concentrations (10 μg/ml and 50 μg/ml).

Conclusions These preliminary results show that a stable ABAT/CD86 interaction may be obtained in cultured ECs, after 24 or 48 hrs from ABAT treatment (100 and 500 μg/ml), resulting in a decreased percentage of CD54+ and CD309+ cells. Although these concentrations are higher than the trough concentrations of abatacept in vivo, they may be physiologically relevant at sites of inflammation such like in RA synovitis.

References

  1. Marrelli A et al. Autoimmun Rev. 2011;10:595-8.

  2. Cutolo M et al. Autoimmun Rev. 2013;12:758-67.

  3. Cutolo M et al. Clin Exp Rheumatol. 2013;31:943-6.

  4. Kreisel D et al. J Immunol 2002;169:6154-61.

Acknowledgements The study was partially supported by research grant (university funds) by Bristol Myers Squibb and the compound for the experiments was provided by Bristol Myers Squibb.

Disclosure of Interest M. Cutolo Grant/research support from: The study was partially supported by research grant (university funds) by Bristol Myers Squibb and the compound for the experiments was provided by Bristol Myers Squibb., S. Soldano: None declared, P. Contini: None declared, A. C. Trombetta: None declared, A. Sulli: None declared, B. Seriolo: None declared, M. A. Cimmino: None declared, S. Paolino: None declared, C. Pizzorni: None declared, P. Montagna: None declared, R. Brizzolara: None declared

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