Biol. Cell (1991), 73

FLOW CYTOMETRY MONITORING OF ORGAN TRANSPLANTATION

BRANDO Bruno, SOMMARUGA Elisabetta. Renal Transplant Unit, Niguarda-Ca' Grands Hospital,

Milano, Italy.

Several flow cytometric (FCM) applications are currently of diagnostic importance in the different phases or organ transplantation (TX). 1) The Dre-TX Crosn Match.

Th i s test discloses the presence of cytotoxic anti-donor antibodies in recipient'a serum. Two different FCM assays are currently available: an indirect jmmunofluorescence test and a c3~tozic one employing propidium iodide. Both show many advantages as compared to conventional'light microscopy techniques. 2) The memsur~ment of immunosunoression.

The complex drug regimens-employed for post-TX immunoauppresaion must be carefully monitored in order to check the correctness of their mechanism of action and to avoid overimmunoauppresaion. The monitor ing of therapeutical i.v. anti-CD3 antibody is of particular importance. 3) The detection of immune events.

Rejection and infection are the most common events after TX, and they still remain difficult to discriminate. Acute viral infections are associated with typical peripheral lymphocyte patterns. In renal TX, CD~.. iymphocytes are sheddedin the urine during acute rejection. 4) ThQ nutnggnn,,nlmntnt:ion of (~D34,,,.eeH~..

The enumerat:.ior~ of circulating hematopoietie precursors is another recent clinical application of FCM. In cancer patients treated with myeloablative doses of chemotherapy and r-Ha GM.CSF, a large number of CD34+ celia can be harvested from peripheral blood and stored to support subsequent courses of therapy with autotransplantation. CD34+ cells are easily and accurately counted by FCM and their number is strictly correlated to their elonogenie properties.

DRCI EXPRESSION IN NORMAL AND PATiiOLOGICAL LYMPtlOID CELLS.

ANTOINE Nadine, LIEINEN Ernst, BOSSELOIR Alain, BECKERS Catherine, SIMAR L~on,

laboratory of Human Histology. University of Liege. rue de Pitteurs. 402 ̂ LIEGE

Since many years, our laboratory focuses its attention on follicular dendritic cells (FDC) in the germinal center of human lymphoid follicles. These cells create a mtcruenvironment necessary for B lymphocytes proliferation and maturation.

In 1983. Naiem et at. produced a munne monoclonai antibody anti- (DRCi) strongly reacting with'human FDC; it has proved of great v',due in the analysis of the distribution of FDC in reacdve lymphoid ttssues and in follicular lymphomas. Some of our results suggest that the DRCl antigen could play a role in the regulation of the intercellular adhesion (Louis et a1.1990). However. this antibody also reacts, though much more weakly, with marginal zone splenic B cells and with some mantle B lymphocytes.

lmmunolabellings performed with ABC complex were not sensitive enough to localize.these cells in tonsillar freeze sections. Although. by flow cytometry, we could reveal DRC! positivity on lymphoid cells populations prepared from human tonsils: 6090 of the B cells appeared DRCI positive. only 10% of T ceils.

Contrary to the tonsillar B cells, blood B cells were DRCI negative.

DRC! expression in lymphoid cells was also found in pathological conditions: EBV transformed mnsillar B ceils and B ceils from chronic lymphocyttc leukemia strongly expressed DRCI.

When the adherence capacity of leukemia cells to isolated FDC was checked, no clear relationship was found between DRCi expression attd adherence. Thus. the function of DRCI remains to be elucidated.

The prognostic value of DRCi antigen expression in leukemias, is under study. REFERENCES: LOUIS E.. PHILIPPET B.. CARDOS B., HEINEN E.. CORMANN N.. KINET-DENOEL C.. BRAUN M. and SIMAR L-J. Acta O.R.L..43, 297- 320. 1990. NAIEM M., GERDES l and ABDULAZIZ, Z. I. Clin. Path.. 36, 167.175, 1983.

50a

DIPYRIDAMOLE INHIBITS THYMIDINE INCORPORATION BUT NOT THE ACQUISITION OF

ACTIVATION MARKERS BY PHA-STIMULATED LYMPHOCYTES

~OUET Nathalie, BROHEE Danv. Lab. Mad. Exp. (ULB), C.H.U. VESALE, Monggny.le-Tilleul.

Dipyridamote (DPM) inhibits thymidine incorporation (3H.Tdr) by PEA.stimulated cells probably by interfering with the membrane transport (1). Others have suggested a real immunodepressive effect (2). This study addresses the effect of DPM on the expression of activation markers by PEA I pM stimulated lymphocytes. Using a FAC Scan flow cytometer, three dual antibody combinations were studied on FicolI-Paque 1,077 isolated and cultured lymphocytes at 0, 24, 48 and 72 h. Ten subjects (30 _- 5 y. old) were investigated. DPM at 10" M concentration abolish 3H.Tdr incorporation but does not affect the expression of HLA.DR (la, clone L243, Becton-Dickinson), IL-2 receptor (clone 2A J, Becton. Dickinson) and CD45R0 (UCHL-I, DAKO) by CD3 (Leu 4) + lymphocytes and lymphoblasts. This experiment confirms the specific effect of DPM on the membrane transport of thymidine without interference with the lymphocyte activation process,

(1) BROHEE D., PIRO P., KENNES B. et NEVE P. Inc J. Immunopbarmac. 8, 925-929, 1986.

(2) BRUSERUD ~. Clin. ImmuaoL Immunopa~oL 42, I02-109, L987.

EVALUATION OF CD4 CD4$RA LYMPHOCYI'ES IN MALIGNANT LYMPHOMA BY CYTOPLUORIMETRY

JACOB Marie.Christine. FAVP~ Minfille and BENSA Jean-Claude Labor~oire d' immJmocytologie, Centre de Transj~sion Sanguine de

Grenoble, BP 33. 38701 !.,4 TRONCHE Codex. FR,4HC£

In malignant diseases, little is known about collaboration between normal T lymphocytes (TIL-T) and tumor cells, In the

present work. we have investigated whether immune T I~mphocytes exist in tumors invaded by B-call non Ha/skin lympoma (B-NHL) or Hodgkin's disease (liD). Therefore, w- have studied the reactivity of the CD45RA monoclonal antibody, which discriminates between naive and memory CD4 T lymphocytes. Our results showed far lower percentages of CD4+ Ci)4$RA+ in malignant lymphoma (30.3% :l: 15,0% in B-HHL, and 37.4% :k 18,6% in liD) than in reactive hypcrplasia (54,7% :I: 13,2%), leading to the conclusion of an accumulation of immune ceUs in tumor micmenvkonmant. A further heterogeneity in the relative proportion of naive and memory TIL-T was al~ observed within lymphoma. In B-NHL, it was related to histological features, as documented by the Kiei classification (p=.028), and to a stonger extend to cytological characteristics attalysed with the Grenoble classification (p<.0001): class 1 NHL, which are essentially indolent NHL displayed lower naive cells (22.2% 4. 7.4%) than class 3 NHL, which are more aggessive (40.1% 4- 16.1%). Among the monoclonal antibodies defining the B cell clone phenotype or activation state (CDI9, CD20, CD21, CD22, CD23, CD24, CD$, CI)I0, CD! la, Ki67), only CD23 (p--.0003) and Ki67 (p=.0007) revealed stati.qical association with the paccntage of naive CD4 iymphocytes. No correlation could be demonstrated with thd proportion of whole TIt-T, activated CD3 DR TILT or CD4 subset.