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B A S I C S C I E N C E

gers a cascade of intracellular events, including the activa-tion of the IL-1-converting enzyme-like cysteine protease(caspase 8), that ultimately leads to nucleoprotein cleavage,DNA fragmentation, and cell apoptosis.6 Fas-FasL interac-tions are involved in the clonal deletion of peripheral T cells,in the downregulation of cytotoxic T-cell activity, and in thespontaneous apoptosis of human neutrophils.6 Thus, Fasseems to be involved in immune downregulation and inpreventing an immune overreaction. It is interesting thatmutations within genes encoding Fas and FasL (lpr and gldmutations, respectively) result in the development of au-toimmune disease in mice.7 Recently, we8 and others9 dem-onstrated that soluble HLA class I (sHLA-I) moleculesupregulate FasL expression and sFasL secretion in activatedFas-positive CD8+ T-cells and trigger their apoptosis. Thelevel of sHLA-I molecules is significantly increased in theserum of patients with an activation of their immune sys-tem, such as occurs during acute rejection of organ al-lografts, acute GVHD after BMT, autoimmune disease, orviral infection.10 Therefore, it has been suggested that sHLA-I molecules may play a significant role in the eliminationof activated T cells after mounting a proper immune re-sponse. If so, serum sHLA-I and sFasL molecules may rep-resent the efferent arm of a network that controls the ex-pansion of CD8+ T-lymphocytes in the course of an immuneresponse.10

It has been reported by other authors11-14 that solublemolecules with immunomodulatory properties are detect-able in commercial albumin, immunoglobulin, and hemo-static preparations. We focused our interest on the detec-tion of sHLA-I and sFasL in various blood components andon the evaluation of their immunomodulatory activities. Wewill briefly summarize the results of our published ex vivostudies15 and report some recent unpublished data fromour laboratory, which suggest that sHLA-I and sFasL mol-ecules may play a major role in transfusion immunomodu-lation.

The concentration of sHLA-I and sFasL molecules inthe supernatant of RBCs stored for 30 days and in plateletswas significantly higher than that in other blood compo-nents, such as RBCs stored for 5 days, prestorage WBC-re-duced RBCs stored for 30 days, washed RBCs, and FFP.15 The

Fas, Fas ligand, and transfusion immunomodulation

Francesco Puppo, Massimo Ghio, Paola Contini, Clemente Mazzei, and Francesco Indiveri

ABBREVIATIONS: FasL = the natural ligand for Fas; MLR(s) =

mixed lymphocyte reaction(s); sFasL = a soluble form of FasL;

SHLA-I = soluble HLA-I.

From the Division of Internal Medicine and Clinical Immunol-

ogy, Department of Internal Medicine, University of Genoa,

Genoa, Italy; and the Blood Bank, Imperia Hospital, Imperia,

Italy.

Address reprint requests to: Francesco Puppo, DiMI, Viale

Benedetto XV n. 6, 16132 Genova, Italy; e-mail:

metclin@unige.it.

Received for publication October 3, 2000; revision received

December 26, 2000, and accepted January 23, 2001.

TRANSFUSION 2001;41:416-418.

Fas (CD95/APO-1) is a 45-kDa type I membrane gly-coprotein that belongs to a family of moleculesthat resemble the receptor for TNF.1 The naturalligand for Fas is called FasL (CD95L), and it is a 40-

kDa type II membrane protein that belongs to the TNF fam-ily.1 Fas has a widespread constitutive tissue expression,being detected in thymus, liver, heart, lung, kidney, andovary.1 In contrast, FasL has a relatively restricted distribu-tion and is predominantly expressed in lymphoid organs aswell as in the stroma cells of the retina and in Sertoli’s cellsin the testes.1 Expression of FasL can be induced in othertypes of cells, such as NK cells, upon exposure to proin-flammatory cytokines. After the activation of peripheralblood T cells, the expression of Fas and FasL is significantlyincreased.2 Activated T cells use a metalloproteinase-likeenzyme to produce a soluble form of FasL (sFasL) consist-ing of a 26-kDa extracellular region of FasL that is releasedwithin microvesicles of 100- to 200-nm diameter.3 This sFasLis detectable in human serum in the course of neoplastic,infectious, and immune-mediated diseases.4 Mature hu-man neutrophils show a constitutive cell-surface coex-pression of Fas and FasL molecules and may release sFasL.5

Fas and FasL are involved in a well-described signal-ing pathway leading to cellular apoptosis.6 Binding of FasLor sFasL to Fas results in receptor aggregation and in theinteraction of a protein called Fas-associated death domainwith the Fas cytoplasmic tail (Fig. 1). This interaction trig-

Fas, FasL, AND TRANSFUSION IMMUNOMODULATION

Volume 41, March 2001 TRANSFUSION 417www.transfusion.org

concentrations of sHLA-I and sFasL molecules correlatedwith the number of residual WBCs and with the length ofstorage. These findings suggest that sHLA-I and sFasL mightderive mainly from residual WBCs that undergo membranechanges during storage.15

The sHLA-I and sFasL molecules in the supernatant ofblood components are functional, and they may exertimmunomodulatory activities. We demonstrated that suchsupernatants blocked antigen-specific cytotoxic T-cell ac-tivity, inhibited the allogeneic mixed lymphocyte reaction(MLR), and induced apoptosis in Fas-positive cells. Theblocking effect on cytotoxic T cells was specifically exertedby sHLA-I molecules, the inhibitory effect on MLRs wasinduced by both sHLA-I and sFasL molecules, andapoptosis was triggered by sFasL molecules.15 Recent un-published data obtained in our laboratory indicate thatimmunomodulatory effects are exerted not only by alloge-neic blood components but also by blood components,which are autologous to the cells utilized in the assays.These results suggest that allogeneic and autologous trans-fusion may exert similar immunomodulatory effects.

Several mechanisms may underlie the immunomodu-latory activities of sHLA-I and sFasL. The sHLA-I moleculesmay bind to CD8 molecules, thus interfering with the rec-ognition of peptides presented in the context of allogeneicand autologous HLA antigens, and they also may upregulatesFasL secretion and apoptosis in activated autologous andallogeneic CD8+ cells. The sFasL molecules may triggerapoptosis in activated Fas-positive T- and B-cells generatedduring immune responses, as well as in neutrophils. More-over, Fas-FasL coexpression and interaction on the mem-brane of activated lymphocytes and neutrophils may causethem to kill themselves (“suicide”) or kill each other (“frat-ricide”) through apoptotic death signaling.

Overall, we suggest that sHLA-I and sFasL moleculesmight play an important role in the proposed immuno-modulatory effects of blood transfusion.16-19 First, thesesoluble molecules suppress the activity and induce theapoptosis of lymphocytes and neutrophils, thereby contrib-uting to the increase in postoperative infection, observedin some studies, among recipients of blood transfusion.Second, they may interfere with immune surveillanceagainst neoplastic cell growth, and therefore they may beinvolved in the suspected increased cancer recurrence intransfused patients. Third, they downregulate immune re-sponses against alloantigens, which may explain, at least inpart, the tolerogenic effect of preoperative blood transfu-sions in some patients receiving allografts.

Research on Fas, FasL, sFasL, and sHLA molecules haswide application in human biology and in the pathogenesisof immune-mediated disorders. Increased understandingof the mechanisms of immune regulation by these mol-ecules will likely result in new insights into the immuneconsequences of allogeneic blood transfusion.

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Fig. 1. Diagram of the mechanism by which FasL triggers cell

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