Structure of the Spleen in Idiopathic Thrombocytopenic Purpura M E H D I TAVASSOLI, M . D . , AND ROBERT M c M l L L A N , M . D . From the L. C. Jacobson Blood Center, Scripps Clinic and Research Foundation, Lajolla,
California
ABSTRACT
of immune-damaged platelets. (Key words: Spleen in thrombocytopenic purpura; Platelets.) IDIOPATHIC
THROMBOCYTOPENIC
PUR-
PURA (ITP) is a clinical and immunologic entity, characterized by thrombocytopenia, shortened platelet survival, and a compensatory increase in thrombopoiesis. 1,6,15 A circulating antiplatelet factor, now generally regarded as an IgG antibody, is present.5'7"9,11,12 Although a similar clinical picture may be present in the course of such disorders as systemic lupus erythemaReceived October 24, 1974; received revised manuscript January 2, 1975; accepted for publication January 2, 1975. Supported by NIH Grants AM-16501 and AM16125 and AEC AT(04-3)899. Dr. Tavassoli is the recipient of a Career Research Development Award 1-K04-AM-70551 from the National Institute of Arthritis, Metabolism and Digestive Diseases. Address reprint requests to Dr. Tavassoli: Scripps Clinic and Research Foundation, 476 Prospect Street, Lajolla, California 92037. 180
tosus and chronic lymphocytic leukemia, idiopathic thrombocytopenic purpura usually occurs de novo. T h e spleen plays a central role in the pathogenesis of this disease, and it has recently been found that this organ is a major site of production of antiplatelet antibody. 12 Circulating platelets are preferentially sequestered by the spleen, 10 wherein an environment suitable for their sensitization exists. Sensitized platelets are then phagocytosed in the cordal compartment of the spleen. Splenectomy is now widely accepted as the therapeutic measure of choice in the management of patients with ITP, resulting in clinical remission in more than 75% of cases.1 Although a few morphologic observations concerning splenic platelet phago-
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Tavassoli, Mehdi, and McMillan, Robert: Structure of the spleen in idiopathic thrombocytopenic purpura. Am J Clin Pathol 64: 180-191, 1975. T h e structure of the spleens of 12 patients with idiopathic thrombocytopenic purpura (ITP) who underwent splenectomy was studied. T h e white pulp was characterized by the presence of large numbers of lymphatic nodules containing highly reactive germinal centers. T h e marginal zone contained large numbers of vessels surrounded by plasma cells. These findings imply active antibody production and are consistent with the concept that the spleen is a major source of antiplatelet antibody production in ITP. Large numbers of platelets in various stages of degradation were seen in cords, particularly in the marginal zone. They appeared both extracellularly and within the cytoplasm of macrophages. It appeared that platelet destruction was initiated by focal cytoplasmic degradation and resulted in formation of cellular debris. These findings indicate that the spleen plays a fundamental role in the pathogenesis of ITP, involving synthesis of antiplatelet antibody, localization of platelets in a milieu rich in this antibody, and subsequently, phagocytosing
August 1975
181
SPLEEN IN ITP
inor
cytosis have appeared in the literature, 3,18 the ultrasturcture of the spleen in I T P has not been extensively investigated. Furthermore, macrophages containing cellular debris have been observed in spleens of patients with thrombocytopenia, 3,18 but no evidence has been presented to indicate the platelet origin of such debris. This study reports the light and electron microscopic findings in spleens from patients with ITP, and places particular emphasis on the pathogenesis of the disease and the fate of platelets within the splenic circulation.
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FIG. 6. Germinal center in ITP. Portion of a large lymphoid cell can be seen (SL). T h e nucleus contains two large nucleoli. T h e cell is reminiscent of PHA-stimulated lymphocytes, x7,500.
with one or two prominent nucleoli (Fig. 6), and are reminiscent of lymphocytes stimulated in vitro with phytohemagglutinin. Macrophages have abundant cytoplasm containing cell debris, the origin of which cannot be recognized (Fig. 5). T h e mantle zone often appears thicker than that of the normal spleen. It contains small lymphocytes, tightly packed in several circumferential shells, set in intervening circumferentially arranged extracellular reticulum (Fig. 3). The marginal zone is distinctive from that of the normal spleen because it contains an unusually large number of vessels, both arteries and sinuses. Large numbers of plasma cells are also present, and tend to aggregate around the small arteries (Fig. 7). Few plasma cells were seen in the marginal zone of our control spleens. Apart from plasma cells, the cellular content of the marginal zone con-
sists of lymphocytes, granulocytes and macrophages (Fig. 7). T h e lymphocytes are similar to those seen in the germinal centers. They are large with relatively abundant cytoplasm and indented nuclei and are distinguished from the small lymphocytes seen in the mantle zone. T h e marginal zone extends into the red pulp, which also contains large numbers of vessels, here mostly sinuses but also few small arteries. Dense globular bodies are frequently encountered in the endothelium of vascular sinuses (Fig. 8). They may appear in secondary lysosomes. The cellular content of the cord consists of granulocytes, platelets, lymphocytes, plasma cells, and a few erythrocytes, but it is particularly rich in macrophages. In spleens from patients 1, 2, and 3, the numbers of granulocytes were also striking. An occasional small aggregate of hemopoietic cells is often seen in this location.
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FIG. 7. Marginal zone in ITP, showing a small artery (A) with several plasma cells (P) in its vicinity. Three granulocytes (G) and three macrophages containing phagocytic materials (arrows) and lymphocytes occupy the rest of this field, x 5,000.
A similar finding was observed in spleens from patients with hereditary spherocytosis and immune hemolytic anemia, but not in the normal spleen. Platelets may be normal in appearance but they show focal degrada-
tion, vesicle formation and degranulation. Macrophages often contain a large amount of cellular debris, some recognizable as partially degraded platelets (Figs. 9 and 10). In PAS-stained preparations, the cytoplasm
August 1975
SPLEEN IN I T P
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FIG. 8. T h e endothelial cell from a splenic sinus occupies most of this field. Note the typically folded nucleus and pinocytic vesicles near the cell membrane. T h e r e are three globular bodies (arrows) in the cytoplasm of this cell, x 11,000.
of these macrophages contain PAS-positive materials (Fig. 4). Platelet Phagocytosis Platelet phagocytosis occurs mainly in the cordal compartment of the red pulp, particularly in the area adjacent to the marginal zone. In this location, platelets are often seen both extracellularly and within the cytoplasm of a macrophage that is tightly applied to the wall of a sinus. Extracellular platelets are sometime normal in appearance, but they may show focal areas of cytoplasmic degradation. Autophagocytosis appears to be a major mechanism of platelet destruction: large numbers of vesicles appear in the cytoplasm, some may contain membranous debris. Some granules appear to consist of membranes without granular content. The platelets located within the cordal macro-
phages often show focal areas of cytoplasmic degradation much more extensive than those seen extracellularly. Parts of platelets become extremely dense, forming debris and sometimes myelin figures, within a morphologically recognizable platelet. Ferritin particles may be seen sprinkled throughout the degrading platelet (Fig. 9). Cellular debris without recognizable platelets is frequently encountered in the cytoplasm of macrophages (Fig. 10). Less frequently, secondary lysosomes containing globular materials are seen in the cytoplasm of both cordal macrophages and the endothelium of vascular sinuses (Figs. 8 and 10). Platelet phagocytosis is not, however, seen in the endothelium of vascular sinuses. Platelet degradation and phagocytosis were particularly conspicuous in spleens from patients 5, 9, and 11, who were considered to be steroid-
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FIG. 9. Most of this field is occupied by a macrophage which is applied to the wall of a sinus. Note the endothelium (END) containing bands of microfilaments (arrow) and the basement membrane (BM) of the sinus. T h e macrophage contains three platelets, identified by numbers. T h e second platelet shows foci of cytoplasmic degradation, indicated by denser areas. T h e first platelet shows a more advanced stage of this process, but, in both, portions of platelet are still recognizable. A third platelet consists entirely of cellular debris. Note the presence of ferritin particles over what may be a phagocytosed structure, x2,200.
resistant, whereas in the spleen of patient 5, considered steroid-responsive, degradation and phagocytosis of platelets were encountered only occasionally.
Comparison with Non-ITP Spleens In the normal spleen and the spleens from patients with HS and AIHA, morphologically normal platelets were fre-
August 1975
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189
quently encountered in the cordal com- which antiplatelet antibody is produced in partment, often in a group, and only rarely response to an antigen as yet unknown, displaying focal areas of cytoplasmic deg- which may be platelet-associated. The antiradation. Somewhat more frequently, dense body is shown to have platelet-binding globular bodies were seen within the sinus specificity.13 T h e antibody-coated platelets endothelium or cordal macrophages, but then undergo degradation and phagocythis finding was not as frequent as in the tosis by macrophages. By virtue of its spleens from patients with ITP. Intra- unique microanatomy, the spleen plays an cellular platelet degradation with forma- important part in every stage of this tion of cellular debris was not found in process. T h e splenic white pulp is an immunologic tissue capable of antigen any of the control spleens. recognition, antigen trapping, and eliciting Discussion an antibody response. 19-20 Previous studies This study emphasizes the central role of in our laboratory have indicated that in the spleen in the pathogenesis of ITP. ITP, the spleen is indeed a major site 13 The disease is an immunologic entity in of production of antiplatelet antibody.
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FlG. 10. T h e cytoplasm of this macrophage contains a large amount of phagocytosed materials that may be platelet-derived. No structure recognizable as a platelet can be seen, however. Note several structures containing globular inclusions, one of which is identified by an arrow. These may correspond to platelet-derived, phospholipid-containing materials seen in light microscopy, x25,000.
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T h e spleen also normally stores large numbers of circulating platelets, permitting them to come into contact with the antibody in an environment where a high concentration of antibody is present. The splenic red pulp is basically a filtration bed 21 where large numbers of macrophages are present. T h e rate of blood flow is slow, thereby permitting adequate time for interaction of the sensitized platelets with cordal macrophages and facilitating platelet phagocytosis.
The PAS-positive macrophages in the lymphatic nodules must be differentiated from similarly stained macrophages in the marginal zone and splenic cords. In the latter location, macrophages often contain
64
platelet debris, and are most frequently seen in those spleens manifesting extensive platelet phagocytosis. Their presence, therefore, does not correlate with the dose of steroids, but with the platelet response to steroids, i.e., they are frequently found in those cases which have not responded to steroid therapy. Platelet destruction in the spleen occurs in the cordal compartment of the red pulp, particularly in the area adjacent to the marginal zone. It may occur in the cytoplasm of macrophages or may begin extracellularly, phagocytosis occurring later during the process. In both situations, the process is initiated by focal degradation of platelets which may be self-perpetuating to result in formation of cellular debris. Cellular debris has previously been found in splenic tissue from patients with thrombocytopenia. 3,8,18 Evidence to indicate the platelet origin of such debris was lacking. The data presented in this study indicate that the cellular debris contained within the macrophages is derived from platelets through the process of focal cytoplasmic degradation. Lipid-containing macrophages, known as "foamy" macrophages, have been observed in the splenic red pulp by several investigators. 3,8,17,18 Histochemical studies indicated that their lipid content consisted chiefly of phospholipids. 17 Because they are frequently seen in states associated with thrombocytopenia, and because platelets have a high content of phospholipids, it has been suggested that the phospholipid content of these macrophages may derive from platelets as a result of platelet phagocytosis. 17 Our observations are in agreement with this concept. Furthermore, the dense globular inclusions we observed in our materials'may correlate with the foamy macrophages described by these investigators, i.e., these inclusions may derive from platelet phagocytosis. However, we observed these inclusions in both cordal macrophages and sinus endothelium. Even
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T h e splenic white pulp in I T P is distinctive because of highly developed secondary lymphatic nodules, 2,16 containing easily recognizable germinal centers, and the presence of large numbers of perivascular plasma cells in the marginal zone. These features indicate splenic activity in antibody production and are consistent with the concept that the spleen is a major site of production of antiplatelet antibody. The "starry-sky" appearance of the germinal centers in hematoxylineosin stained preparations probably results from cell death and phagocytosis by macrophages. These macrophages contain large amounts of phagocytic inclusions and cell debris corresponding to the "tingible" bodies discussed by Fliedner. 4 These inclusions may appear as PASpositive materials in preparations stained with PAS-hematoxylin. Lymphocytolysis, caused by steroid therapy for thrombocytopenia, appears to accentuate the "starry-sky" appearance and increases the number of macrophages containing PASpositive materials. Thus, these macrophages were seen most frequently in those spleens from patients who had received the most extensive steroid therapy, irrespective of the platelet response to steroids.
A.J.C.P.—Vol.
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here, the inclusion can derive from platelets. T h e presence of platelets in the sinus endothelium has been described, 14 and it has been suggested that this endothelium, capable of phagocytosis and pinocytosis but lacking the necessary lysosomal machinery, may selectively take u p platelets to use their lysosomes for processing the materials it has pinocytized. 14 T h e platelet remnants then may be seen in sinus endothelium as the inclusions described herein. References
bocytopenia and other blood dyscrasias treated with steroids. Am J Clin Pathol 39:607-615, 1963 9. Hirschman RJ, Shulman NR: T h e use of platelet serotonin release as a sensitive method for detecting antiplatelet antibodies and a plasma antiplatelet factor in patients with idiopathic thrombocytopenic purpura. Br J Haematol 24:793-802, 1973 10. Jandl J H , Aster RH: Increased splenic pooling and the pathogenesis of hypersplenism. Am J Med Sci 253:383-397, 1967 11. Karpatkin S, Siskind GW: In vitro detection of platelet antibody in patients with idiopathic thrombocytopenic purpura and systemic lupus erythematosus. Blood 33:795-812, 1969 12. McMillan R, Longmire RL, Yelenosky R, et al: Immunoglobulin synthesis in vitro by splenic tissue in idiopathic thrombocytopenic purpura. N Engl J Med 286:681-684, 1972 13. McMillan R, Longmire RL, Yelenosky R, et al: T h e quantitation of platelet-binding IgG produced in vitro by spleens from patients with idiopathic thrombocytopenic purpura. N Engl J Med 291:812-817, 1974 14. Molnar Z, Rappaport H: Fine structure of the red pulp of the spleen in hereditary spherocytosis. Blood 39:81-98, 1972 15. Najean Y, Ardaillou N, Dresch C, et al: T h e platelet destruction site in thrombocytopenic purpuras. Br J Haematol 13:409426, 1967 16. Nickerson DA, Sunderland DA: T h e histopathology of idiopathic thrombocytopenic purpura hemorrhagica. Am J Pathol 13:463490, 1937 17. Saltzstein S: Phospholipid accumulation in histiocytes of splenic pulp associated with thrombocytopenic purpura. Blood 18:73-88, 1961 18. Summerall JM, Gibbs WN: Splenic histiocytosis associated with thrombocytopenia. Acta Haematol (Basel) 48:34-38, 1972 19. Weiss L: T h e white pulp of spleen. Bull Johns Hopkins Hosp 115:99-174, 1964 20. Weiss L: T h e Cells and Tissues of the Immune System. Englewood Cliffs, N.J., Prentice Hall, 1972, pp 4 9 - 7 1 21. Weiss L, Tavassoli M: Anatomical hazards to the passage of erythrocytes through the spleen. Semin Hematol 7:372-380, 1970
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1. Baldini M: Idiopathic thrombocytopenic purpura. N Engl J Med 274:1245-1251, 1 3 0 1 1306, 1360-1367, 1966 2. Bowman HE, Pettit VD, Caldwell FT, et al: Morphology of the spleen in idiopathic thrombocytopenic purpura. Lab Invest 4 : 2 0 6 216, 1955 3. Firkin BG, Wright R, Miller S, et al: Splenic macrophages in thrombocytopenia. Blood 33:240-245, 1969 4. Fliedner TM: On the origin of tingible bodies in germinal centers, Germinal Centers in Immune Responses. Edited by NO Cottier $$, $$, et al. New York, Springer-Verlag, 1967, p 218 5. Handin RI, Piessens WF, Moloney WC: Stimulation of non-immunized lymphocytes by platelet antibody complexes in idiopathic thrombocytopenic purpura. N Engl J Med 289:714-718, 1973 6. Harker LA: Thrombokinetics in idiopathic thrombocytopenic purpura. Br J Haematol 19:95-104, 1970 7. Harrington WJ, Sprague CC, Minnich V, et al: Immunologic mechanisms in idiopathic and neonatal thrombocytopenic purpura. Ann Intern Med 38:433-469, 1953 8. Hill JM, Speer RJ, Gedikogln H: Secondary lipoidosis of spleen associated with throm-
191
California
ABSTRACT
of immune-damaged platelets. (Key words: Spleen in thrombocytopenic purpura; Platelets.) IDIOPATHIC
THROMBOCYTOPENIC
PUR-
PURA (ITP) is a clinical and immunologic entity, characterized by thrombocytopenia, shortened platelet survival, and a compensatory increase in thrombopoiesis. 1,6,15 A circulating antiplatelet factor, now generally regarded as an IgG antibody, is present.5'7"9,11,12 Although a similar clinical picture may be present in the course of such disorders as systemic lupus erythemaReceived October 24, 1974; received revised manuscript January 2, 1975; accepted for publication January 2, 1975. Supported by NIH Grants AM-16501 and AM16125 and AEC AT(04-3)899. Dr. Tavassoli is the recipient of a Career Research Development Award 1-K04-AM-70551 from the National Institute of Arthritis, Metabolism and Digestive Diseases. Address reprint requests to Dr. Tavassoli: Scripps Clinic and Research Foundation, 476 Prospect Street, Lajolla, California 92037. 180
tosus and chronic lymphocytic leukemia, idiopathic thrombocytopenic purpura usually occurs de novo. T h e spleen plays a central role in the pathogenesis of this disease, and it has recently been found that this organ is a major site of production of antiplatelet antibody. 12 Circulating platelets are preferentially sequestered by the spleen, 10 wherein an environment suitable for their sensitization exists. Sensitized platelets are then phagocytosed in the cordal compartment of the spleen. Splenectomy is now widely accepted as the therapeutic measure of choice in the management of patients with ITP, resulting in clinical remission in more than 75% of cases.1 Although a few morphologic observations concerning splenic platelet phago-
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
Tavassoli, Mehdi, and McMillan, Robert: Structure of the spleen in idiopathic thrombocytopenic purpura. Am J Clin Pathol 64: 180-191, 1975. T h e structure of the spleens of 12 patients with idiopathic thrombocytopenic purpura (ITP) who underwent splenectomy was studied. T h e white pulp was characterized by the presence of large numbers of lymphatic nodules containing highly reactive germinal centers. T h e marginal zone contained large numbers of vessels surrounded by plasma cells. These findings imply active antibody production and are consistent with the concept that the spleen is a major source of antiplatelet antibody production in ITP. Large numbers of platelets in various stages of degradation were seen in cords, particularly in the marginal zone. They appeared both extracellularly and within the cytoplasm of macrophages. It appeared that platelet destruction was initiated by focal cytoplasmic degradation and resulted in formation of cellular debris. These findings indicate that the spleen plays a fundamental role in the pathogenesis of ITP, involving synthesis of antiplatelet antibody, localization of platelets in a milieu rich in this antibody, and subsequently, phagocytosing
August 1975
181
SPLEEN IN ITP
inor
cytosis have appeared in the literature, 3,18 the ultrasturcture of the spleen in I T P has not been extensively investigated. Furthermore, macrophages containing cellular debris have been observed in spleens of patients with thrombocytopenia, 3,18 but no evidence has been presented to indicate the platelet origin of such debris. This study reports the light and electron microscopic findings in spleens from patients with ITP, and places particular emphasis on the pathogenesis of the disease and the fate of platelets within the splenic circulation.
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FIG. 6. Germinal center in ITP. Portion of a large lymphoid cell can be seen (SL). T h e nucleus contains two large nucleoli. T h e cell is reminiscent of PHA-stimulated lymphocytes, x7,500.
with one or two prominent nucleoli (Fig. 6), and are reminiscent of lymphocytes stimulated in vitro with phytohemagglutinin. Macrophages have abundant cytoplasm containing cell debris, the origin of which cannot be recognized (Fig. 5). T h e mantle zone often appears thicker than that of the normal spleen. It contains small lymphocytes, tightly packed in several circumferential shells, set in intervening circumferentially arranged extracellular reticulum (Fig. 3). The marginal zone is distinctive from that of the normal spleen because it contains an unusually large number of vessels, both arteries and sinuses. Large numbers of plasma cells are also present, and tend to aggregate around the small arteries (Fig. 7). Few plasma cells were seen in the marginal zone of our control spleens. Apart from plasma cells, the cellular content of the marginal zone con-
sists of lymphocytes, granulocytes and macrophages (Fig. 7). T h e lymphocytes are similar to those seen in the germinal centers. They are large with relatively abundant cytoplasm and indented nuclei and are distinguished from the small lymphocytes seen in the mantle zone. T h e marginal zone extends into the red pulp, which also contains large numbers of vessels, here mostly sinuses but also few small arteries. Dense globular bodies are frequently encountered in the endothelium of vascular sinuses (Fig. 8). They may appear in secondary lysosomes. The cellular content of the cord consists of granulocytes, platelets, lymphocytes, plasma cells, and a few erythrocytes, but it is particularly rich in macrophages. In spleens from patients 1, 2, and 3, the numbers of granulocytes were also striking. An occasional small aggregate of hemopoietic cells is often seen in this location.
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186
TAVASSOLI AND MCMILLAN
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FIG. 7. Marginal zone in ITP, showing a small artery (A) with several plasma cells (P) in its vicinity. Three granulocytes (G) and three macrophages containing phagocytic materials (arrows) and lymphocytes occupy the rest of this field, x 5,000.
A similar finding was observed in spleens from patients with hereditary spherocytosis and immune hemolytic anemia, but not in the normal spleen. Platelets may be normal in appearance but they show focal degrada-
tion, vesicle formation and degranulation. Macrophages often contain a large amount of cellular debris, some recognizable as partially degraded platelets (Figs. 9 and 10). In PAS-stained preparations, the cytoplasm
August 1975
SPLEEN IN I T P
•
*
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FIG. 8. T h e endothelial cell from a splenic sinus occupies most of this field. Note the typically folded nucleus and pinocytic vesicles near the cell membrane. T h e r e are three globular bodies (arrows) in the cytoplasm of this cell, x 11,000.
of these macrophages contain PAS-positive materials (Fig. 4). Platelet Phagocytosis Platelet phagocytosis occurs mainly in the cordal compartment of the red pulp, particularly in the area adjacent to the marginal zone. In this location, platelets are often seen both extracellularly and within the cytoplasm of a macrophage that is tightly applied to the wall of a sinus. Extracellular platelets are sometime normal in appearance, but they may show focal areas of cytoplasmic degradation. Autophagocytosis appears to be a major mechanism of platelet destruction: large numbers of vesicles appear in the cytoplasm, some may contain membranous debris. Some granules appear to consist of membranes without granular content. The platelets located within the cordal macro-
phages often show focal areas of cytoplasmic degradation much more extensive than those seen extracellularly. Parts of platelets become extremely dense, forming debris and sometimes myelin figures, within a morphologically recognizable platelet. Ferritin particles may be seen sprinkled throughout the degrading platelet (Fig. 9). Cellular debris without recognizable platelets is frequently encountered in the cytoplasm of macrophages (Fig. 10). Less frequently, secondary lysosomes containing globular materials are seen in the cytoplasm of both cordal macrophages and the endothelium of vascular sinuses (Figs. 8 and 10). Platelet phagocytosis is not, however, seen in the endothelium of vascular sinuses. Platelet degradation and phagocytosis were particularly conspicuous in spleens from patients 5, 9, and 11, who were considered to be steroid-
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FIG. 9. Most of this field is occupied by a macrophage which is applied to the wall of a sinus. Note the endothelium (END) containing bands of microfilaments (arrow) and the basement membrane (BM) of the sinus. T h e macrophage contains three platelets, identified by numbers. T h e second platelet shows foci of cytoplasmic degradation, indicated by denser areas. T h e first platelet shows a more advanced stage of this process, but, in both, portions of platelet are still recognizable. A third platelet consists entirely of cellular debris. Note the presence of ferritin particles over what may be a phagocytosed structure, x2,200.
resistant, whereas in the spleen of patient 5, considered steroid-responsive, degradation and phagocytosis of platelets were encountered only occasionally.
Comparison with Non-ITP Spleens In the normal spleen and the spleens from patients with HS and AIHA, morphologically normal platelets were fre-
August 1975
SPLEEN IN ITP
189
quently encountered in the cordal com- which antiplatelet antibody is produced in partment, often in a group, and only rarely response to an antigen as yet unknown, displaying focal areas of cytoplasmic deg- which may be platelet-associated. The antiradation. Somewhat more frequently, dense body is shown to have platelet-binding globular bodies were seen within the sinus specificity.13 T h e antibody-coated platelets endothelium or cordal macrophages, but then undergo degradation and phagocythis finding was not as frequent as in the tosis by macrophages. By virtue of its spleens from patients with ITP. Intra- unique microanatomy, the spleen plays an cellular platelet degradation with forma- important part in every stage of this tion of cellular debris was not found in process. T h e splenic white pulp is an immunologic tissue capable of antigen any of the control spleens. recognition, antigen trapping, and eliciting Discussion an antibody response. 19-20 Previous studies This study emphasizes the central role of in our laboratory have indicated that in the spleen in the pathogenesis of ITP. ITP, the spleen is indeed a major site 13 The disease is an immunologic entity in of production of antiplatelet antibody.
Downloaded from http://ajcp.oxfordjournals.org/ by guest on June 6, 2016
FlG. 10. T h e cytoplasm of this macrophage contains a large amount of phagocytosed materials that may be platelet-derived. No structure recognizable as a platelet can be seen, however. Note several structures containing globular inclusions, one of which is identified by an arrow. These may correspond to platelet-derived, phospholipid-containing materials seen in light microscopy, x25,000.
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TAVASSOLI AND MCMILLAN
T h e spleen also normally stores large numbers of circulating platelets, permitting them to come into contact with the antibody in an environment where a high concentration of antibody is present. The splenic red pulp is basically a filtration bed 21 where large numbers of macrophages are present. T h e rate of blood flow is slow, thereby permitting adequate time for interaction of the sensitized platelets with cordal macrophages and facilitating platelet phagocytosis.
The PAS-positive macrophages in the lymphatic nodules must be differentiated from similarly stained macrophages in the marginal zone and splenic cords. In the latter location, macrophages often contain
64
platelet debris, and are most frequently seen in those spleens manifesting extensive platelet phagocytosis. Their presence, therefore, does not correlate with the dose of steroids, but with the platelet response to steroids, i.e., they are frequently found in those cases which have not responded to steroid therapy. Platelet destruction in the spleen occurs in the cordal compartment of the red pulp, particularly in the area adjacent to the marginal zone. It may occur in the cytoplasm of macrophages or may begin extracellularly, phagocytosis occurring later during the process. In both situations, the process is initiated by focal degradation of platelets which may be self-perpetuating to result in formation of cellular debris. Cellular debris has previously been found in splenic tissue from patients with thrombocytopenia. 3,8,18 Evidence to indicate the platelet origin of such debris was lacking. The data presented in this study indicate that the cellular debris contained within the macrophages is derived from platelets through the process of focal cytoplasmic degradation. Lipid-containing macrophages, known as "foamy" macrophages, have been observed in the splenic red pulp by several investigators. 3,8,17,18 Histochemical studies indicated that their lipid content consisted chiefly of phospholipids. 17 Because they are frequently seen in states associated with thrombocytopenia, and because platelets have a high content of phospholipids, it has been suggested that the phospholipid content of these macrophages may derive from platelets as a result of platelet phagocytosis. 17 Our observations are in agreement with this concept. Furthermore, the dense globular inclusions we observed in our materials'may correlate with the foamy macrophages described by these investigators, i.e., these inclusions may derive from platelet phagocytosis. However, we observed these inclusions in both cordal macrophages and sinus endothelium. Even
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T h e splenic white pulp in I T P is distinctive because of highly developed secondary lymphatic nodules, 2,16 containing easily recognizable germinal centers, and the presence of large numbers of perivascular plasma cells in the marginal zone. These features indicate splenic activity in antibody production and are consistent with the concept that the spleen is a major site of production of antiplatelet antibody. The "starry-sky" appearance of the germinal centers in hematoxylineosin stained preparations probably results from cell death and phagocytosis by macrophages. These macrophages contain large amounts of phagocytic inclusions and cell debris corresponding to the "tingible" bodies discussed by Fliedner. 4 These inclusions may appear as PASpositive materials in preparations stained with PAS-hematoxylin. Lymphocytolysis, caused by steroid therapy for thrombocytopenia, appears to accentuate the "starry-sky" appearance and increases the number of macrophages containing PASpositive materials. Thus, these macrophages were seen most frequently in those spleens from patients who had received the most extensive steroid therapy, irrespective of the platelet response to steroids.
A.J.C.P.—Vol.
August 1975
SPLEEN IN I T P
here, the inclusion can derive from platelets. T h e presence of platelets in the sinus endothelium has been described, 14 and it has been suggested that this endothelium, capable of phagocytosis and pinocytosis but lacking the necessary lysosomal machinery, may selectively take u p platelets to use their lysosomes for processing the materials it has pinocytized. 14 T h e platelet remnants then may be seen in sinus endothelium as the inclusions described herein. References
bocytopenia and other blood dyscrasias treated with steroids. Am J Clin Pathol 39:607-615, 1963 9. Hirschman RJ, Shulman NR: T h e use of platelet serotonin release as a sensitive method for detecting antiplatelet antibodies and a plasma antiplatelet factor in patients with idiopathic thrombocytopenic purpura. Br J Haematol 24:793-802, 1973 10. Jandl J H , Aster RH: Increased splenic pooling and the pathogenesis of hypersplenism. Am J Med Sci 253:383-397, 1967 11. Karpatkin S, Siskind GW: In vitro detection of platelet antibody in patients with idiopathic thrombocytopenic purpura and systemic lupus erythematosus. Blood 33:795-812, 1969 12. McMillan R, Longmire RL, Yelenosky R, et al: Immunoglobulin synthesis in vitro by splenic tissue in idiopathic thrombocytopenic purpura. N Engl J Med 286:681-684, 1972 13. McMillan R, Longmire RL, Yelenosky R, et al: T h e quantitation of platelet-binding IgG produced in vitro by spleens from patients with idiopathic thrombocytopenic purpura. N Engl J Med 291:812-817, 1974 14. Molnar Z, Rappaport H: Fine structure of the red pulp of the spleen in hereditary spherocytosis. Blood 39:81-98, 1972 15. Najean Y, Ardaillou N, Dresch C, et al: T h e platelet destruction site in thrombocytopenic purpuras. Br J Haematol 13:409426, 1967 16. Nickerson DA, Sunderland DA: T h e histopathology of idiopathic thrombocytopenic purpura hemorrhagica. Am J Pathol 13:463490, 1937 17. Saltzstein S: Phospholipid accumulation in histiocytes of splenic pulp associated with thrombocytopenic purpura. Blood 18:73-88, 1961 18. Summerall JM, Gibbs WN: Splenic histiocytosis associated with thrombocytopenia. Acta Haematol (Basel) 48:34-38, 1972 19. Weiss L: T h e white pulp of spleen. Bull Johns Hopkins Hosp 115:99-174, 1964 20. Weiss L: T h e Cells and Tissues of the Immune System. Englewood Cliffs, N.J., Prentice Hall, 1972, pp 4 9 - 7 1 21. Weiss L, Tavassoli M: Anatomical hazards to the passage of erythrocytes through the spleen. Semin Hematol 7:372-380, 1970
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1. Baldini M: Idiopathic thrombocytopenic purpura. N Engl J Med 274:1245-1251, 1 3 0 1 1306, 1360-1367, 1966 2. Bowman HE, Pettit VD, Caldwell FT, et al: Morphology of the spleen in idiopathic thrombocytopenic purpura. Lab Invest 4 : 2 0 6 216, 1955 3. Firkin BG, Wright R, Miller S, et al: Splenic macrophages in thrombocytopenia. Blood 33:240-245, 1969 4. Fliedner TM: On the origin of tingible bodies in germinal centers, Germinal Centers in Immune Responses. Edited by NO Cottier $$, $$, et al. New York, Springer-Verlag, 1967, p 218 5. Handin RI, Piessens WF, Moloney WC: Stimulation of non-immunized lymphocytes by platelet antibody complexes in idiopathic thrombocytopenic purpura. N Engl J Med 289:714-718, 1973 6. Harker LA: Thrombokinetics in idiopathic thrombocytopenic purpura. Br J Haematol 19:95-104, 1970 7. Harrington WJ, Sprague CC, Minnich V, et al: Immunologic mechanisms in idiopathic and neonatal thrombocytopenic purpura. Ann Intern Med 38:433-469, 1953 8. Hill JM, Speer RJ, Gedikogln H: Secondary lipoidosis of spleen associated with throm-
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