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EOSINOPHILIC DROPLET FORMATION, A MORPHOLOGIC MANIFESTATION OF CARDIOMYOCYTE APOPTOSIS, IS PRESENT IN THE HUMORAL (MICROVASCULAR) REJECTION OF HUMAN CARDIAC ALLOGRAFTS

1998; Wolters Kluwer; Volume: 66; Issue: 3 Linguagem: Inglês

10.1097/00007890-199808150-00023

1534-6080

Jiri T. Beranek,

Mechanical Circulatory Support Devices

The recent observation in Transplantation(1) that eosinophilic droplet (ED) formation represents a morphologic expression of apoptosis, the main mechanism of cardiomyocyte death in hyperacute rejection, raises immediately the question of whether the same phenomenon is not present in other forms of cardiac rejection. Lones et al. (2) have described clinicopathological features of cardiac "humoral" rejection in 81 consecutive patients. Its histopathology included capillary endothelial cell swelling, the obstruction of capillaries by swollen macrophages, immunoglobulin and complement deposition in microvasculature, interstitial edema and hemorrhage, and neutrophil infiltration. Since ED formation has often been mistaken for interstitial hemorrhage, the latter must be re-evaluated for the presence of the former. In Figure 2A of the Lones article (2), numerous cardiomyocytes manifest "contraction band necrosis" (3). Here and there, the contraction bands fuse, forming extended areas of hypereosinophilic hyalinized tissue (Fig. 2, A, B, and C). In some places, this hyalinized tissue fragments into particles from which the smallest correspond to EDs and not to red cells (Fig. 2, A, B, and C). In Figure 2E (2), a biopsy specimen "in which interstitial hemorrhage is more pronounced" is presented. Its analysis shows, however, that the alleged red cells do not originate in the interstitial space but appear by the fragmentation of cardiomyocytes. This visual evidence is further substantiated by the following facts: (a) Some EDs within cardiomyocytes manifest the longitudinal striation which is in continuity with the longitudinal striation of the myocardial cell, indicating their common derivation. (b) Cell nuclei fragments are present inside of some EDs, excluding the possibility that these are red cells. (c) Not all EDs display the hypereosinophilic color of red cells. Their color may vary from the deep pink of myofibers to the bright red of red cells, suggesting their myocardial origin. (d) The cardiomyocytes situated in the vicinity of the alleged interstitial hemorrhage do not manifest any compression or any deviation of their long axes, revealing that a substantial amount of contractile tissue must have been eliminated, presumably, by fragmenting into EDs. (e) EDs are of unequal size and cannot, therefore, be red cells. The above morphologic analysis is not to deny a presence of the concomitant genuine interstitial hemorrhage resulting from the immunological damage of microvasculature in humoral rejection (2,4). Moreover, ED formation itself creates contractile tissue defects in which the interstitium is no longer supported by cardiomyocytes and may undergo a disruption resulting in interstitial hemorrhage. Thus, the similarity between ED formation and interstitial hemorrhage and their spatial and temporal closeness have contributed to their confusion. As a result, ED formation has been overlooked. This fact is best demonstrated in hemorrhagic infarctions (5,6). In the article by Fujiwara et al. (6), the fragmentation of myofibers into EDs is clearly discernible in Figure 2C, questionable in Figures 2B and 2A1, and most probably already absent from Figure 2A2. Clinically, cardiac humoral rejection is characterized by hemodynamic compromise usually explained by the immunological damage of microvasculature resulting in ischemia of the irrigated contractile tissue (2,4). Further sequence of events from ischemia to apoptosis is not known, but two hypotheses, both involving damage of the sarcolemma, may be envisaged. According to Hack et al. (7), ischemic cells are not able to maintain phospholipid asymmetry among the inner and the outer leaflets of the normal cell membrane, the outer leaflet containing mainly sphingomyelin and phosphatidylcholine (PC) and the inner leaflet, phosphatidylserine (PS) and phosphatidylethanolamine (PE). In ischemia, the phospholipids of the inner and the outer leaflets undergo "flip-flop" exchange, which results in the increased amount of PS and PE in the outer leaflet. Flip-flopped membranes are sensitive to hydrolysis by secretory phospholipase A2, which generates lyso-PC in the outer leaflet. This compound binds circulating acute phase C-reactive protein (C-RP). Human C-RP bound to ischemic cardiomyocytes fixes circulating complement subcomponent C1q and thereby activates the classical complement pathway, generating factors needed for the clearance of pathological tissue(8). Among these, terminal component-complex is of particular importance because of its lytic capabilities. It forms pores in the sarcolemma of primed cardiomyocytes, and in this way, permits the entry of the factor that initiates apoptosis into the cell(1). ED formation is a widespread phenomenon in cardiac pathology(9). If provoked experimentally, it may appear with extraordinary rapidity (1,10,11) in previously normal animals with extremely low C-RP serum levels(8). This precludes an effective interaction between C-RP and complement in the pathogenesis of cardiomyocyte apoptosis, and another explanation for the increased permeability of the sarcolemma must be found. ED formation also appears in transmyocardial laser revascularization when a suitable low level of laser energy that does not provoke accidental cell death is used (12-14). In these cases, it is easy to imagine that laser energy itself has damaged the sarcolemma and allowed the entry of a trigger of apoptosis into the cell. Similarly, total ischemia in infarction experiments also injures the sarcolemma effectively (3), leading to a massive ED formation after reperfusion (10). All of these data suggest that cardiomyocyte apoptosis is triggered by a physiological substance easily available in the interstitial space. Extracellular Ca2+ is about 10,000 times more concentrated than free ionized calcium in cytosol (15). Any damage of the sarcolemma permits, therefore, its flux into the cell. There, it activates calcium-sensitive protease calpain, known for its involvement in the apoptosis of other cells (1). This makes the influx of extracellular ionized calcium the most suitable candidate for triggering cardiomyocyte apoptosis, regardless of whether the pore-forming action of complement has been or has not been involved. In complex clinical situations, such as the humoral rejection of cardiac allografts, both the interaction between C-RP and complement and the direct damage of the sarcolemma by ischemia may take place. Jiri T. Beranek Columbia, Missouri 65203