The TAR syndrome is a rare eongenital disorder eharaeter ized by the absenee of radb on both arms and severe throm boeytopenia during the first years of life. Reeent studies showed that the thromboeytopenia seen in TAR patients is eaused by a defeet in the megakaryocytopoiesis/thromboey topoiesis.^4~7 Different possible meehanisms for this bone marrow failure ean he postulated: (1) absenee of humoral or eellular stimulators of megakaryoeytopoiesis, (2) absenee of megakaryoeytie progenitor eells, (3) eellular defeets in megakaryoeytie preeursors (eg, reeeptor defeets), or (4) pres

Fig 1. Expressicn of c-MpI on piatelets from TAR patients. IAI c-MpI expression on the surface of plateiets es measured by flow cytometry see Materials and Methodsl. Grav. isotype Gontrol; black. anti c-Mpi. IB Detection of c-Mpl in pletelet lysates after immunoprecipitetion and Western blot analysis with anti-c.Mpl entibody Ml lchemo luminescence detection, See Meterjeis and Meth odsl. Lene 1, heelthv donor; lene 2, TAR patient na. 1.

enee of humoral or edlular inhibitors of megakaryoeyto poiesis.

There are eontroversial reports eonceming Mega-CSA or “TPO-like" aetivities in the sera of TAR patients. Our group⁶ and others⁴ found high Mega-CSA in the sera of TAR patients. However, de Alareon et a1⁷ reported Mega-CSA and “TPO-like" activities in the serum of one TAR patient. whieh were within the range of normal and below the high levels seen in amegakaryoeytie subjects. Miehaleviez et a1⁵ even found an inhibitory effeet of plasma of one TAR patient on the growth of multipotent and megakaryoeytie progeni tors. After the diseovery of TPO and its reeeptor e-Mpl we were able to measure the aetivity of this major humoral regulator of megakaryoeytopoiesis and thrombocytopoiesis in the sera of TAR patients. We found elevated levels of bioaetive TPO in sera from all TAR patients tested, exelud ing a TPO produetion defeet as the eause of thrombocyto penia in TAR syndrome. lt is a eommon phenomenon that

lineage speeifle hematopoietic growth factors are inversely regulated with the number of eorresponding mature eells in peripheral blood. This has been shown for G-CSF,²⁸ EPO.^29,30. and TPO^31,32 and was found to be the ease in a number of eongenital eytopenias as the severe eongenital neutropenia. also.³³ The first determination of TPO serum levels of patient 2 yielded no elevated level of this eytoldne. We have no explanation of that diserepancy. However, the first determi nation has been made with a relatively old serum sample and we eannot exelude a loss of activity during inappropriate storage. On the other hand, variations of TPO levels in TAR syndrome might be the reason for differenees in former re ports.

The presence of eolony forming units megakaryoeyte (CFU-Mega) in the bone marrow of TAR patients is another pomt of eontroversy in the literature. Whereas most groups, like us, did not find any growth of megakat'yocytic eolonies from TAR patients.⁴'⁶ de Alareon et a1⁷ showed normal CFU  

the c-Mpl on the platelets from the TAR patients was found tu be 86 kD as determined bv SDS-PAGE and was not different from that of healthy controls (Fig 1B).

Costiruulotjou of p/atelets with TPO. TPO svnereizes with ADP or thrombin receptor agonists to activate platelets as measured by increased expression of P-seleetin (CD62P(.² We used this fact for in vitro testing of TPO reaetivitv of platelets from TAR patients. (CD62P).˛7 served as an aetivation-dependent marker. Using this method, platelets from normal individuals showed a synergism of rhTPO (5 - 10 ng/mL) with the platelet aetivators ADP and TRAP. AI though there was a great variety in the reacüvity of platelets tu the activators ADP and TRAP, platelets from all TAR patienis showed reactivity in the range of normal controls (Fig 2 and Table 4). However, we eould not deteet the ex peeted synergism of these platelet aetivators with rhTPO up to enneentrations of 1 µg/mL in these patients. The results of one representative expetiment are shuwn in Fig 2. To quantitate the synergistic effeet of TPO on platelet activation we ealeulated the ratio between the amount of CD62P-posi tive eells afier TPO costimulation (with ADP or TRAP) and the amount of CD62P-positive eells after control stimulation without TPO (Table 4). TPO preineubation (20 ng/mL) lead tu a l.6-fold (~0.3) enhaneement of ADP stimulation (50 µmol/L) or 1 4-fold (~0.3) enhaneement of TRAP stimula tion (5 µmol/L) in platelets of normal individuals (n = 6). In enutrast, TPO stimulation indiees in TAR patients were

1.0 0.1 for ADP and 1.0 0.2 for TRAP. respeetively

(Table 4).