Norihiko Shimoyama

Expression of p16 protein and hypermethylation status of its promoter gene in adenoid cystic carcinoma of the head and neck

Reduced expression of the p16 gene product (protein), an inhibitor of cyclin-D-dependent protein kinase which regulates cell cycle at the G1/S boundary, is implicated in tumor progression in various neoplasms. Hypermethylation of the p16 promoter gene has recently been suggested to be one of the reasons for the reduced protein expression. To explore the role of p16 in the biological behavior of adenoid cystic carcinomas (ACC), we investigated the immunohistochemical expression of p16 protein in 38 ACC tumors (32 primary, 3 recurrent, and 3 metastatic tumors) and the methylation status of its promoter gene. We also examined their relationships to the histological grade of malignancy. Positive reaction of p16 protein was demonstrated in the nuclei of luminar cuboidal cells in areas with tubular patterns. The reactions were reduced in the areas with solid or large cribriform patterns. The levels of p16 expression correlated with the histological grade of malignancy. Recurrent or metastatic tumors did not differ with respect to histological grades from the original tumor except for 1 case, in which p16 expression was reduced compared to the primary tumor. Methylation-specific PCR demonstrated the hypermethylation status of the p16 promoter gene in 4 of 22 primary tumors (21%), all of which showed negative or low expression of the p16 protein. The study indicated that p16 expression was reduced in ACC cases of higher histological grade of malignancy and that hypermethylation of its promoter gene may be involved in its process in some cases.

Pathohistological and immunohistoehemieal studies on Castleman's disease of the lymph node

Histological and immunohistochemical examinations were carried out on lymph nodes in 9 cases with Castlemans disease. In all cases, there were whorled follicle centers and proliferation of blood vessels. These findings were also considered to be important criteria for diagnosis of the plasma cell type of this disease. The whorled follicle centers often consisted of smaller concentrical structures and contained factor VIII-positive cells. In some cases, there were many small whorled structures surrounding postcapillary venules within the interfollicular areas of lymph nodes. These structures had factor VIII-positive granules indicating their endothelial origin. In 2 cases of the plasma cell type, monoclonal plasma cell proliferation (IgA, λ and IgG, λ, respectively) and cellular atypia were identified. These findings showed that extramedullary plasmacytoma could occur not only in the multicentric form, but also in the solitary form of Castlemans disease. In the lymph node of a case of the hyaline-vascular type, there were scattered tumour nodules consisting mainly of factor VIII-positive, atypical spindle cells suggesting associated Kaposis sarcoma. It is conceivable that an abnormal immune state plays a role in pathogenesis of Castlemans disease.

GANGLIONEUROBLASTOMA CONTAINING SEVERAL KINDS OF NEURONAL PEPTIDES WITH WATERY DIARRHEA SYNDROME

This report presents an adrenal ganglioneuroblastoma containing several kinds of neuronal peptides. The tumour was found in an autopsy case of a 3‐year‐old girl with clinical manifestation of intractable diarrhea, hypokalemia, achlorhydria, and with elevated levels of plasma vasoactive intestinal peptide (VIP). Immunoperoxidase staining showed many immuno‐reactive VIP‐ containing cells, some somatostatin‐and substance P‐contalning cells on the tumour sections. Ultrastructurally, the tumour cells contained numerous secretory granules that could be divided mainly into two types; one is a small cored vesicle (50‐150 nm in diameter) and the other large electron dense secretory granule (200‐500 nm in diameter). It was suggested that the cells in ganglioneuroblastoma derived from neural crest are closely related to the cells that could differentiate into gut‐hormone‐producing cells.

PRIMARY CEREBRAL MALIGNANT LYMPHOMA WITH MACROGLOBULINEMIA AND CARCINOMA OF THE COLON

A case of primary cerebral malignant lymphoma, immunoblastic type was reported. The immunoelectrophoresis of the patient serum showed an abnormal bow of IgM lambda type. Autopsy revealed that the cerebral lymphoma showed no extracranial spread except for adenocarcinoma of the colon with hepatic metastasis. Ultrastracturally, some of the lymphoma cells showed plasmacytoid differentiation. Immunoperoxidase study showed intracytoplasmic polyclonal immunoglobulins in non‐neoplastic plasma cells around the colonic carcinoma, in the bone marrows, and spleen. However, neither IgM nor lambda light chain was found in the cerebral lymphoma cells. The source of the macroglobulinemia in this case was discussed.

Peroxidase-positive acute leukemia with T-cell markers

Recently, Jehn et al. reported leukemic cells with Auer-bodies in case of acute lymphocytic leukemia (ALL) [1], whereas definite positive peroxidase (POX) reactions in the majority of the leukemic cells in patients suffering from ALL has not yet been described. We here present the first case of an acute leukemia in which the neoplastic cells bear simultaneously T-cell characters defined by immunologic and cytochemical studies and myeloid characters by positive POX reactions. The patient, a 64-year-old man, was diagnosed as suffering from acute leukemia in February of 1980. At that time the total white blood cell count was 27.5 x 109/1, of which more than 90% were atypical lymphoblastoid cells, and the bone marrow was almost replaced by these cells. The 10-20/~m large leukemic cells had round or oval nuclei with one to two prominent nucleoli and a narrow cytoplasm with occasional vacuoles. Some of these cells showed a few azurophilic granules. Electron microscopically these cells displayed the structures of leukemic lymphoblasts. Rosette analysis of the leukemic cells, separated by density gradient, gave the following results; E: 20% and EAC:0%. In a complement-dependent cytotoxicity test 88 % of the separated cells reacted with highly specific anti-human T-lymphocyte serum (ATS) [2]. ATS absorbed with these cells was no longer cytotoxic to normal human peripheral T-cells. Surface and intracytoplasmic immunoglobulins were negative. POX activity was demonstrated using benzidine and carbazole as substrates. The percentage of leukemic cells with positive reactions was 85 and 63, respectively. Coarse granular reaction pattern was observed. 3-Amino-I, 2,4-triazole had no effect on these reactions while sodium acide almost completely inhibited them. Positive POX reactions were also found on many E-rosette forming cells in cytocentrifuged smears. Electron-microscopic examination for POX confirmed these positive reactions. On the other hand, more than 60% of the cells showed strong focal reactions for acid phosphatase and acid esterase, which have been generally accepted as characteristics of T cells. About half of the leukemic cells revealed fine granular reactions for PAS.

Systemic amyloidosis complicated with peliosis.

Dear Editor, A 75-year-old male was hospitalized in April 2006 for oral and urinary bleeding. He presented with hematoma in the left latissimus dorsi and with hepatomegaly, splenomegaly, anemia, and thrombocytopenia. His platelet count was 49× 10/μL, his prothrombin time was 15.2 s, his FDP was 196.5 μg/dL, and his fibrinogen was 121 mg/dL. His serum haptoglobin level was reduced but autoantibody and Coombs tests were negative. Serum vitamin B12 and folic acid levels were normal. These findings suggested the presence of disseminated intravascular coagulation (DIC). Computed tomography (CT) revealed hepatomegaly, splenomegaly, and several 10-mm areas of low density in the spleen (Fig. 1a). In magnetic resonance imaging (MRI) these low-density areas appeared as high-density areas in T1 and T2 images (Fig. 1b, c) but were not enhanced in the dynamic phase (Fig. 1d). We thought that the splenomegaly was complicated by hematoma, possibly related to chronic DIC. We could not identify the mechanism of this hemolysis and DIC. After prednisolone (0.5 mg/kg) was administered daily because autoimmune hemolytic anemia could not be ruled out, the anemia and thrombocytopenia decreased and the patient’s bleeding tendency decreased enough that the rate of transfusion could be reduced. Splenonectomy on 9 June 2006 revealed that the spleen weighed 175 g and contained many blood-filled spaces located perifollicularly (Fig. 2a). The spleen showed the ‘bacon spleen’ appearance characteristic of amyloidosis. The appearance of the lining of these spaces differed from that of the endothelium. Amorphous eosinophilic materials observed in the red pulp of the spleen (Fig. 2b) were positive for direct fast scarlet (DFS) stain and periodic acid Schiff (PAS) stains and were immunohistochemically positive for amyloid P component (Fig. 2c, d). The pathological diagnosis was peliosis. The anemia, thrombocytopenia, and hematoma of the left latissimus dorsi abated after the splenonectomy. Although the thrombocytopenia abated, the patient sometimes passed a bloody stool after the prednisolone was discontinued. Gastrointestinalscopy (GIS) and colonoscopy (CS) at the time of the melena revealed nothing abnormal. The patient was discharged from hospital in August 2006, but his bleeding tendency continued and in October 2006 he had a hemorrhage in the left femoris muscle. These symptoms abated after administration of carbazochrome sodium sulfonate and tranexamic acid, but the hepatomegaly gradually became more aggravated. In April 2007 he was readmitted because of uncontrollable ascites caused by portal hypertension due to the hepatomegaly. Administration of potassium canrenoate, furosemide, and albumin did not reduce the ascites, nor did a peritoneovenous shunt. The ascites was accompanied by pleural effusion, and both gradually increased until the Ann Hematol (2009) 88:917–920 DOI 10.1007/s00277-008-0690-6

PRECURSOR T CELL LEUKEMIA

A case of acute lymphoblastic leukemia with a giant splenomegaly in an 8‐year‐old boy was investigated by immunological, cytochemical and electronmicroscopical techniques. Bone marrow and peripheral blood were largely replaced by large blast cells with a nonconvoluted nucleus. Cytochemically, most of the blast cells showed strong focal acid phosphatase activity. In the surface marker studies, these blast cells formed EAC rosettes but not E rosettes, while they showed positive reactivity with anti‐T lymphocyte serum but not with anti‐B lymphocyte serum. This membrane phenotype E‐, C3R+, T+, B‐ suggested that leukemic blast cells in this patient presumably originated from precursor T cells.