In accordance with the revised criteria of FAB classification in 1985 (Ann Intern Med 10) by FAB Co-operative Group, 32 cases diagnosed as erythroleukemia (EL) were re-classified. They were examined for the abnormal morphological findings of erythroblasts and FAB classification was re-evaluated for its validity.
Eleven cases belonged to M6 (Group A), while 21 cases did not belong to M6 (Group B). Control groups included 7 hematologically normal cases, 8 cases of megaloblastic anemia and 6 cases of hemolytic anemia.
Ultrastructural observation on erythroblasts from all cases was done and the presence of the abnormal findings was examined. Erythroblasts in control groups were also analysed because abnormal findings were found. A comparison of Group A and Group B was made by means of frequency of abnormal findings.
Statistically, atypism of the cytoplasm was marked in Group A, whereas atypism of the nucleus was marked in Group B. A correlation of abnormal findings between Group A and Group B suggented that atypism of the calls of Group A was more remarkable than Group B and discriminant function revealed the possibility of classification into two groups.
The FAB classification of optical examination corresponds with that of the ultrastructural findings, and this suggests an adequacy of revised classification.

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Ultrastructual features of a human erythroidcell line (YN-1) were studied before and fater induction of erythroid differentiation. YN-1 was estabsished by us from a patient with Ph¹ chromosome-positive chronic myelocytic leukemia who had undergone an erythroblastic transformation. YN-1 cells lacked the membrane characteristics of lymphocytes and myelomonocytic cells and carried on their surface glycophorin A. Analysis of the hemoglobin by isoelectric focusing indicated major bands in the region of embryonic hemoglobin. Erythroid differentiation of YN-1 cells was induced by hemin, cytosine arabinoside and actinomycin D. When they were cultured in the liguid media during exponential growth without inducer of erythroid differentiation, most cells were circular or oval with a few microvilli. The nucleus of them were circular showing the euchromatic pattern with a few prominent nucleoli. Numerous polysomes were dispersed throughout the cytoplasm. A small number of mitochondria and undeveloped Golgi apparatus were observed. A few micropinocytic vesicles and siderosomes were seen. After induction of erythroid differntiation by hemin, YN-1 cells had a low nuclear hytoplasmic ratio and exhibited aggregation of nuclear chromatin. Organelles such as mitochondria, Golgi apparatus and siderosome were developed and increasing number of micropinucytic vesicles were seen. Increased cytoplasmic electron density was observed, and it was correlated with the strength of pseudeperoxidase in DAB procedure, indicating hemoglobin synthesis.
From these results, relation of functional differentiation and morphological differentiation was discussed.

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Bone marrow materials from twenty-seven cases with myelodysplastic syndrome (MDS) before receiving any therapy which contained 5 of refractory anemia (RA), 4 of RA with ring sideroblast (RARS), 8 of RA with excess of blasts (RAEB), 3 of RAEB in transformation (RAEB-T), and 7 of chronic myelomonocytic leukemia (CMMoL) were studied. Bone marrow specimens from these cases were examined by transmission electron microscopy combined with myeloperoxidase (MPO) cytochemistry. Dyserythropoiesis in ultrastructural level was defined as follows: nuclear clefts, nuclear blebs, abnormal division of nucleus, irregular contour of nucleus, degenerative changes of nucleus, dilated nuclear envelope, iron diposits of mitochondria, large or degenerative changes of mitochondria, siderosomes and megaloblastic changes.
The morphological analysis revealed the presence of nuclear clefts and blebs to be a sign of poor prognosis (p<0.01), namely these patients died within one year because of infections and/or bleedings. But there was no significant difference between dyserythropoiesis and the tendency to leukemic transformation.
Dyserythropoiesis were occasionally seen in some patients with aplastic anemia, pernicious anemia, hemolytic anemia, myeloproldferative disorders and so on. The erythroblasts in acute erythremia were completely different from these of MDS. At the present time, we concluded in ultrastructural study that dysplastic changes in erythroblasts are important value for diagnosis of MDS but they can not to be a marker for acute leukemia.

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Erythroblasts in patients with liver cirrhosis were electron microscopically studied by vertically cut sections of the buffy coat of the aspirated bone marrow, sideroblasts distributed from the top to the bottom of which were evaluated as a percentage of all erythroblasts present. 12 patients, six of whom were alcoholic, had percentages of abnormal sideroblasts with iron deposited in the mitochondria ranging from 5% to 80%, in whom, however, counts of ringed sideroblasts in ordinary smear specimens with Prussian blue staining were not accounted for up to 1%. Namely, the intramitochondrial iron studies in sideroblasts of cirrhotics have produced conflicting results. This difference is understood to result from that the intramitochondrial iron deposits are toosmall for resolution limit ability of the light microscope and optically invisible. This optically invisible iron probably plays to diminish ringed sideroblast ratios. All of non-alcoholic patients detected abnormal sideroblasts had anemia (6 cases). Despite no anemia, positive cases of abnormal sideroblasts were all alcoholic (4 cases). Correlation of red cell indices, serum iron, total iron binding capacity, folic acid or ferritin, and the recognition of abnormal sideroblasts was not seen. Although ringed sideroblasts are scanty in number or absent and further studies are needed, this sideroblast seems to represent a morphologic evidence of iron metabolic abnormality in cirrhotics.

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In order to disclose the relation between function and fine structural morphology, important components of the erythroblasts, such as Fe, hemoglobin, RNA and glycoconjugates, were observed by using ultrastructural cytochemical techniques.
Dialysed iron method for acid mucopolysaccharides stained nuclear envelope as well as cell coat in mature eryththroblasts. The positivity of nuclear envelope was not observed in other types of bone marrow cells, suggesting that denucleation of the erythroblasts needed cytochemical equivalence in nuclear envelope and plasma membrane.
The fine structural distribution of periodate-reactive glycoconjugates was observed with the periodic acid-thiocarbohydrazide-silver proteinate (PA-TCH-SP) method. Glycogen particles, lysosomes and Golgi apparatus were PA-TCH-SP-positive in nomal erythroblasts Glycogen particles in erythroblasts were remarkably increased in diseases exhibiting dyserythropoiesis such as RAEB, erythroleukemia, transient abnomal myelopoiesis of Down's syndrome and dyserythropoiesis of juvenile chronic mylocytic leukemia. In diseases showing hypererythropoiesis such as iron deficiency anemia, very small premature infants, an increase in glycogen was observed in most cases.
Iron, the main component for synthesizing hemoglolin, is detected as electron dense materials by standard EM morphology, However, when the iron is small quantity, it is difficult to observe by the standard EM morphology. In this case, prussian blue staining of electron microscope is useful to desify the feruginous materials.
The fine structural distribution of homglobin was observed by using the antihemoglobin-protein A-gold technique, The gold particles, which indicated the distribution of hemoglobin, were observed in the cytoplasm and nucleus. The concentration of the gold particles increased with the maturation of the erythroblasts, and maximum in mature red blood cells. Intranuclear hemoglobin was increased in erythroblasts from congesital dyserythropoietic anemia due to influx of hemoglobin from the cytoplasm through enlarged nuclear pores.
The RNase-gold complex method was applied for the distribution of RNA in erythroblasts Young erythroblasts exhibited heavy deposit of the gold suggesting that RNA synthesis was active before the start of hemoglobin synthesis. The Golgi area, granules and mitochondria lacked the gold particles The deposition of the gold was decreased as maturation progressed, and no RNA reaction was detected in mature red blood cells.

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INTRODUCTION: β-Thalassemia is a relatively rare congenital disorder characterized by hemolysis and ineffective erythropoiesis due to impaired synthesis of globin β-chain, while CDAs are other types of rare disorders defined by peculiar nuclear abnormality of erythroblasts. We experienced an interesting case with features of both β-thalassemia and CDA-I.
CASE REPORT: A 27-yr old male was referred to Tenri Hospital for the evaluation of anemia. He was diagnosed as having “aplastic anemia” at 4 yr of age. At 20 yr of age, the patient was diagnosed as having β-thalassemia. His liver and spleen were enlarged and each was palpable 3 cm below the costal margin. The hematological findings were as follows: Hb 4.2g/dl, Retics 0.8%, PLT 9.9×10⁴/cu mm, WBC 2100/cu mm. The red cell morphology showed anisopoikilocytosis, anisochromia, tear drop cells, leptocytes and target cells. Bone marrow examination revealed hyperplasia of erythroid precursors with megaloblastoid change and multinuclearity. Anisocytosis and abnormal nuclear segmentation were observed in megakaryocytes, and imbalanced maturation of nucleus and cytoplasm in myeloid precursors. Blood chemistry showed normal range of serum VB₁₂, VB₆ and folic acid. CFU-GM and CFU-E were also within normal range. Ferrokinetic study showed shortened PIDT1/2, enhanced PIT and markedly reduced % RCU. Half life of ⁵¹Cr-labeled RBC was shortened (14.9 d). The data described above indicate that the cause of the patient's anemia was a combination of ineffective erythropoiesis and hemolysis. No abnormal Hb was observed, although Hb F (2,3%) and Hb A₂ (5.3%) were increased The ratio of β-chain synthesis to α-chain synthesis was 0.58, indicating that this case was the heterozygote of β-thalassemia. Electron microscopic study showed various morphological abnormalities of bone marrow erythroblasts; partial loss of nuclear membrane, spongy and donut-like apperrance of heterochromatin, and nuclear clefts (CDA-I); aggregated dense granules in the nucleus and loop-like protrusion of the cytoplasm (thalassemia); iron-laden mitochondria and autophagolysosomes (both CDA-I & thalassemia).
DISCUSSION: Unexpectedly severe anemia in this heterozygous thalassemia patient probably resulted from the complication of CDA-like nuclear abnormalities Ultrastructural study of erythroblasts in anemic patients of unknown cause may be helpful in revealing the cause of anemia.

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Immunological abnormalities have been reported in myelodysplastic syndromes (MDS), but little is known about the incidence of positive direct Coombs' test in MDS. The immunoglobulin levels and autoantibody profiles of 29 patients with primary MDS, classified according to the FAB criteria, were analysed. Of the 29 patients, 11 patients (37.9%) showed a polyclonal rise in serum immunoglobulins and four patients (13.8%) had low immunoglobulin levels. Out of 23 patients tested, the direct Coombs' test was positive in three (13.0%). Of these three patients, a polyclonal rise in serum immunoglobulins was present in two, rheumatoid factor in one, and anti-DNA antibody in two. Two of these patients had autoimmune hemolytic anemia. We report these patients with positive direct Coombs' test in MDS, and also discuss a correlation between immunological abnormalities and their clinical course.

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Between 1972 and 1985, 21 children less than 1 year of age were treated with treatment programs of the Children's Cancer & Leukemia Study Group for acute lymphoblastic leukemia (ALL).
The presenting features and natural history were assesed in these infants with ALL (group I). The results were compared with that of the other two groups (Group II: 483 children between the ages of 1 and 9, Group III: 83 children over than 10 years).
The infants with ALL presented with a higher prevalence of leukocyte counts over 100×10³/mm³, hepatosplenomegaly, thrombocytopenie and shorter duration of remission than those in the other two groups.
The continuous complete remission rate of these infants with ALL was 43.0% at 1 year.
Some infections complication during induction and maintenance phase were experienced. Two patients had several septic episodes. Two patients had syndrome of inappropriate scecretion of antidiuretic hormone the other patients had chicken pox.
The prognosis of ALL is especially poor when the diagnosis were before 1 year of age. It can only be improved by a very intensive chemotherapeutic approach, which, particularly at this age, implies equally intensive and specific supportive cares. Also, alternative approaches to central nervous system prophylaxis are needed to reduce the high prevence of CNS disease and toxicity.

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A case of adult T-cell leukemia (ATL) presenting with pancytopenia and comlicating diabetes insipidus was reported. A 40-year old man was admitted to our hospital because of easy fatigability in Feburary 1985. He was physically unremarkable except for cervical and inguinal lymphadenopathy and moderate hepatomegaly. Blood examination revealed Hb 5.9g/dl, platelets 20,000/mm³, and leukocytes 2,400/mm³ with 33% abnormal lymphocytes showing convoluted or lobulated nuclei. Bone marrow histologically showed significant hemopoietic depletion and scattering foci of abnormal lymphocytes without fibrosis. Lymph node biopsy showed malignant lymphoma, diffuse, medium cell type. T cell subset analysis showed cells from peripheralblood and lymph node to have helper/inducer phenotype. Anti-ATLA antibody was positive. From these findings, he was diagnosed as ATL. In late April 1985, polydipsia, polyuria (>10L/day) with a specific gravity of 1004-1008 appeared. Brain CT showed suprasellar mass. CSF cytology was positive for leukemic cells. The water-depression test and an administration of vasopressin suggested central diabestes insipidus. He died of pulmonary complication 3 months after onset of diabetes insipidus although chemotherapy and radiation therapy resulted in diminution of suprasellar mass and reduction of urine volume. Autopsy revealed necrosis of the infundibulum without leukemic infiltlation In addition, peripheral blood and lymph node mononuclear cells and serum did not suppress either normal erythroid or granuloid colony-formation.

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A 39 year-old male was admitted to our hospital with a complaint of macrohematuria. Cystoscopical examination revealed a prominent lesion at bladder neck. Hemotological examination showed marked thrombocytopenia. The bone marrow aspiration on admission revealed marked infiltration of lymphoblastic cells, which had deeply basophilic cytoplasm and a few small vacuoles. At this time, diagnosis of L2 was made according to FAB classification. Pathological finding of biopsied bladder mucosa indicated an invasion of leukemic cells. Leukemic blasts obtained from bone marrow aspiration were surface immunoglobulin positive and Ia positive. Cytogenetic analyses showed specific translocation, t (8; 14) (q24, q32). According to all these findings, this patient can be classified as Burkitt's type ALL (FAB L3). He was treated with VEMP therapy and complete remission was achieved after all. But relapsed after 2 weeks of first remission Additional chemothemapy was not effective at all and he died from sepsis 4 months after the diagnosis.
In this case, we clearly demonstrated an importance of immunological and cytogenetic examinations for detailed diagnosis of ALL. They play an important role for the prediction of prognosis.

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We report a case of Ph¹-positive acute lymphoblastic leukemia whose blast cells were biphenotypic and bigenotypic.
He was diagnosed as having ALL with Ph¹ chromosome. Surfase markers were investigated, and B₄, J₅, My₇, and My₉, were shown to be positive J₅ and My₉ were simultaneously expressed on the same blast cells by two-color flowcytometry. Gene analysis was made and rearrangement of heavy chain of immunoglobulin gene and β-chain of T cell receptor gene was observed Although the origin of the blast cells was not clarified, we think that the cells were so immature that they had abilities to differentiate to several lineages of T cells, B cells, and myeloid cells, or that the immature cells were disturbed to differentiate due to malignant transformation and they remained at an early stage.
He was treated with doxorubicin, vincristine, and predonisolone, and obtained complete remission.

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We report here that treatment with cyclosporine induced remission in a severe case of 48-year-old female, pure cell aplasia. In this patient, treatment with prednisolone and oxymetholone responded better temporarily after thymectomy, but hemoglobin value gradually decreased during 2 years. Treatment with prednisolone plus azathioprine, fluoxymesterone, or nandrolone decanoate was unsatisfactory and multiple blood transfusions were repeated for three and a half years. Two hundred mg of cyclosporine was started from June, 1986. A striking reticulocytosis peaking around 4 weeks was seen and there was a rapid increase in hemoglobin levels 5.0 to 12.9 g/dl and remission has been maintained for over four and a half months.
T lymphocyte mediated suppression of erythropoiesis was suggested by the result of in vitro erythroid colony study (CFU-E) with or without patient's lymphocytes and the effect of cyclosporine. The treatment with cyclosporine might be recommended in such cases.

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Spontaneous rupture of the spleen is a rare clinical condition. Two cases are presented in association with non-Hodgkin lymphoma of different histologic types, namely, diffuse: mixed type and diffuse: large cell type (clear cell variant).
In the first case the ruptured spleen was discovered at autopsy. In the second case, however, the patient developed acute abdomen at the initial stage of the disease and the emergency laparotomy disclosed the rupture of the spleen.
The mechanism of spontaneous splenic rupture was discussed with the review of previously reported cases. Though some infarcted areas were observed in both of the ruptured spleens, the true cause of spontaneous rupture is still unknown

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A 59-year-old man was admitted in December 1985, because of cough, edema, fever, eruption, generalized lymphadenopathy and ascites. Laboratory examination showed leukocytosis, severe anemia, thrombocytopenia and abnormality of coagulation Bone marrow aspirate was hypocellular, especially erythroblasts were rarely observed. Immunological studies showed polyclonal hypergammaglobulinemia, positive Coombs' test and anti-DNA antibody. A lymph node biopsy specimen exhibited the typical histologic picture of immunoblastic lymphadenopathy (IBL)-like T cell lymphoma. The patient was treated with prednisolone and showed marked symptomatic improvement. Platelet counts and abnormality of coagulation normalized with the improvement of lymphadenopathy and hypergammaglobulinemia However, a recovery from the impaired erythropoiesis was delayed He died of pneumonia in February 1986. IBL-like T cell lymphoma is a rare disease and a variaty of immunological abnormalities was reported. Our case showed various clinical pictures with hypoplastic anemia, thrombocytopenia and abnormality of coagulation, which could be based on the autoimmune mechanisms.

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A 62-year old woman complained of fever, general fatigue and child's fist-sized subcutaneous masses. Lymphocytosis from 12,400 to 22,000/μl had been observed, for five months before admission. On admission, a plain X-ray film disclosed a tumor shadow in the right upper pulmonary field. Neither hepatosplenomegaly nor rashes were found. White blood cell count in peripheral blood was 19,700/μl with lymphocytosis of 68%. Peripheral lymphocytes formed rosettes with sheep erythrocytes and OKT3-, OKT4- and OKT10-positive lymphocytes were markedly increased in number, but OKT8- and IL-2R-positive ones were decreased. Anti-ATLA antibody and ATLV proviral DNA were not detected. The biopsy tissue obtained from the subcutaneous mass of the left upper arm showed a histological appearance of diffuse lymphoma of the large cell type in the Japanese LSG classification. At autopsy, tumors were found in the subcutis of the cervix, the right upper lobe of the lung, the submucosa of the esophagus and the paraaortic lymph nodes. Electron microscopic examination suggested that tumor cells and leukemic cells were of the same cell line. The tumor cells were considered to be transformed from the leukemic cells.

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