Combined fibrolamellar carcinoma and cholangiocarcinoma exhibiting biphenotypic antigen expression: a case report.

Fibrolamellar carcinoma (FLC), a variant of hepatocellular carcinoma (HCC), very rarely occurs in association with cholangiocarcinoma (CC). This report describes the first case of FLC coexisting with CC (FLC-CC) from Japan. Although the major part of the tumour located in the right lobe of the liver showed the typical features of FLC, CC was admixed with the FLC, not only in the primary hepatic tumour, but also in the lymph node metastases. Immunohistochemical analysis revealed that, although carcinoembryonic antigen (CEA), which can be detected with monoclonal antibodies in the cytoplasm and the cell surface of CC cells but not HCC cells, was expressed in only the CC cells in the primary tumour, it was expressed extensively in the cytoplasm of both CC and FLC cells in the metastatic and recurrent tumours. Furthermore, Hep Par 1, a hepatocyte specific antigen, was also expressed in both the FLC and CC cells. These findings suggest that, in this case, both FLC and CC were possibly derived from the same cancer stem cell with the capacity to differentiate into both hepatocytes and bile duct epithelium, and that both the cellular components, therefore, exhibited biphenotypic antigen expression.

Identification of novel direct transcriptional targets of glucocorticoid receptor.

Transcription of the genes Granzyme A (GZMA), FK506 binding protein 51 (FKBP5), and Down syndrome critical region gene 1 (DSCR1) is upregulated in leukemic cells upon treatment with glucocorticoids (GCs). Several lines of evidence suggest that these genes are implicated in GC-induced apoptosis upstream of the Bcl-2 family of proteins. These genes were upregulated by GC even in the presence of an inhibitor of protein synthesis, cycloheximide, indicating that they are direct target genes of glucocorticoid receptors. DSCR1 is reported to have four isoforms, each of which has a distinct first exon, E1-E4. Among these isoforms, the one with E1 was selectively upregulated by GC. GZMA and FKBP5 have a cluster of putative glucocorticoid response elements (GREs) in introns 1 and 2, respectively, that was identified to be responsible for the response to GC. They were composed of one complete (A/T)G(A/T)(A/T)C(A/T) sequence surrounded by two incomplete (A/T)G(A/T)(A/T)C(A/T) sequences separated by one to four nucleotides. DSCR1, however, did not have a functional GRE upstream or downstream of exon 1. These studies may lead to improved therapeutic uses of GCs in leukemia and lymphoma based upon the expression of these GC target genes.

Study to increase the frequency of autopsies performed for cases of infant deaths--proposed revision of the law on postmortem examination and corpse preservation and other related regulations.

By definition, sudden infant death syndrome (SIDS) requires diagnosis through exclusion by conducting an autopsy. To obtain a reliable diagnosis of this disease, an autopsy is essential. However, the frequency with which autopsies are conducted in Japan is not sufficient to meet the need associated with the diagnosis of SIDS. To improve this frequency, various public policies, such as nationwide implementation of the administrative autopsy system (medical examiner system), the application of the practice of autopsy approved by families, and legally required autopsies, are being considered; but none has been put into practice. On the other hand, attention has been called to the fact that the Law on postmortem examination and corpse preservation, which was instituted at the end of the Second World War, requires updating. In the current report, it is proposed that the following be added to Article 8, item 3 of this Law: "the Metropolitan or Prefectural Governor must insist that an autopsy be conducted on all cases of a sudden and unexpected death of an infant to investigate the cause of this death." At present, the annual incidence of SIDS in Japan is reported to be 500. To put the above-recommended legal requirement into practice, the estimated annual addition to the budget, if conducted as approved or an administrative autopsy, will be in the order of 150,000-500,000 dollar, which is within the prescribed limits for an appropriation.

Frequent increase of DNA copy number in the 2q24 chromosomal region and its association with a poor clinical outcome in hepatoblastoma: cytogenetic and comparative genomic hybridization analysis.

In a cytogenetic and comparative genomic hybridization (CGH) study of 38 hepatoblastomas, we found gain of 1q in 17 tumors (44.7%), that of 2 / 2q in 14 (36.8%), that of 20 / 20q in 9 (23.7%) and that of 8 / 8q in 8 (21.0%), loss of 4q in 4 (10.5%) and no DNA copy changes with normal karyotype or no mitotic cells in 11 (28.9%). Eleven tumors with 2 / 2q gain detected by CGH had a total chromosome 2 gain, a partial 2q gain, or a total chromosome 2 gain with an augmented partial 2q region; the common region for DNA copy gain was 2q24. Two-color fluorescence in situ hybridization (FISH) analyses using probes covering the centromere of chromosome 2 or HOXD13 (2q31) confirmed the CGH findings, and showed that the common region for gain in 2q was centromeric to HOXD13. Event-free survival (EFS) +/- standard error (SE) at 5 years was lowest in patients with 2q gain [37 +/- 15%], highest in those with no DNA copy changes [82 +/- 12%], and intermediate in those with DNA copy changes other than 2q gain [74 +/- 13%] (P = 0.0549). Multivariate analysis showed that 2q gain was an independent factor predicting a poor outcome. These findings suggest the presence of a growth-promoting gene or an oncogene in the 2q24 chromosome band, and a tumor suppressor gene in terminal 4q, which have important roles in the development and progression of hepatoblastoma.

Quantitative analysis of hypertrophy in cardiac chambers in cyanotic tetralogy of Fallot.

Although early total corrective repair for cyanotic tetralogy of Fallot is now safely performed at many institutions, long-term complications after surgical repair have been demonstrated. Therefore, the optimal procedure and timing for surgical treatment remain controxersial. In the present study, we conducted a quantitative analysis of the hypertrophy of all four chambers of 87 autopsied hearts of cyanotic tetralogy of Fallot and 71 normal control hearts utilizing the myocardial mass index, and evaluated the progression of lesions with advancing age. In cyanotic tetralogy of Fallot, hypertrophy of the right ventricle progresses immediately after birth, with that of the right atrium developing soon after. The left side of the heart is normal or slightly atrophied which could be corrected by sufficient palliative intervention or total corrective repair. The growth curves of both ventricles were parallel to those of normal hearts for the period studied. Pulmonary atresia, palliative operation, and total corrective repair have been shown to have some influence on the morphological characteristics of hearts of cyanotic tetralogy of Fallot.

The developmental and aging changes of Down's syndrome cell adhesion molecule expression in normal and Down's syndrome brains.

We studied the expression of Down's syndrome cell adhesion molecule (DSCAM) in Down's syndrome (DS) and control brains, using antisera against peptide fragments of DSCAM. On Western blots of human, mouse and rat brain homogenates, the antisera recognized a product at approximately 200 kDa. In the brain of a 2-year-old patient with DS, Western blotting revealed an overexpression of DSCAM compared to an age-matched control. Immunohistochemistry demonstrated DSCAM in the cerebral and cerebellar white matter of both control and DS subjects, in accordance with the temporal and spatial sequence of myelination. In DS brains, immunoreactivity for DSCAM, compared to that for controls, was enhanced in the Purkinje cells at all ages, and in the cortical neurons during adulthood. In demented DS patients, DSCAM immunoreactivity was observed in the core and periphery of senile plaques. The pattern of DSCAM expression suggests that it may play a role as an adhesion molecule regulating myelination. The overexpression of DSCAM may also play a role in the mental retardation and the precocious dementia of DS patients, although the mechanism of neuronal dysfunction is undetermined.

Differentiation-associated expression and intracellular localization of cyclin-dependent kinase inhibitor p27KIP1 and c-Jun co-activator JAB1 in neuroblastoma.

Neuroblastoma is a unique pediatric cancer, which spontaneously regress in some infants and undergo maturation in older children. The cyclin-dependent kinase inhibitor p27KIP1 negatively control cell cycle progression and its expression is reported to be associated with differentiation and prognosis of some human cancers. To examine whether p27KIP1 is involved in differentiation of neuroblastomas, expression and localization of p27KIP1 in 30 cases of neuroblastic tumors were determined with immunohistochemistry. p27KIP1 was expressed in all cases, but staining intensity and intracellular localization varied in association with tumor differentiation. Primitive small round neuroblasts showed negative or only weak nuclear staining, while differentiating tumor cells displayed a novel, intense cytoplasmic positivity besides the nuclear staining, and mature ganglion cells showed intense positive reaction confined to the nucleus. A neuroblastoma cell line TGW was also immunostained positively for p27KIP1 in the cytoplasm after differentiation induction, and western blot analysis revealed an increase of p27KIP1 in these cells, corroborating the in vivo observations. JAB1, which is thought to bind p27KIP1 and transport it from the nucleus to the cytoplasm for proteasome/ubiquitin-mediated degradation, was found to be localized both in the cytoplasm and the nucleus in undifferentiated and differentiating tumors whereas located predominantly in the nucleus of differentiated tumor cells. These data indicate that the cytoplasmic localization of p27KIP1 in the process of differentiation is due to upregulation of p27KIP1 synthesis and subsequent degradation and suggest a role of p27KIP1 in differentiation of neuroblastoma.

Pancreatoblastoma. Three original cases and review of the literature.

PURPOSE: To describe the characteristics of pancreatoblastoma. MATERIAL AND METHODS: We studied 3 cases of pancretoblastoma and reviewed another 59 cases. Parameters analyzed were tumor site, hemorrhage, capsule formation, necrosis, vascularity, production of alpha-fetoprotein (AFP), cystic changes and calcification. RESULTS: The diagnostic findings were as follows: pancreatic head origin (24/54, 44%), pancreatic body and tail origin (30/54, 56%), hemorrhage (16/17, 94%), capsule formation (24/26, 92%), necrosis (28/31, 90%), hypervascularity (10/14, 71%), production of AFP (19/28, 68%), cystic changes (11/16, 69%), and calcification (10/21, 48%). All neonatal cases demonstrated cystic changes. Three of them were patients with Beckwith-Wiedmann syndrome. The incidence of capsule formation and calcification was not related to the origin of the tumor. CONCLUSION: The most common features of pancreatoblastoma are hemorrhage, capsule formation and necrosis.

Tritium in [15O]water, its identification and removal.

The present investigation was undertaken to identify the long-lived radionuclide and its chemical forms existing in [15O]water which was synthesized from 15O produced by the nuclear reaction 14N(d,n)15O, and to develop a method for its removal to facilitate radioactive waste disposal. The long-lived nuclide was identified as tritium based on a comparison of its physical half-life and the energy spectrum of beta-rays with those of tritium. The major chemical form of tritium in the target gas was estimated to be molecular hydrogen. The tritium radioactivity was completely removed without a serious loss occurring to the yield of [15O]water by passing the irradiated target gas over a heated palladium catalyst followed by a calcium chloride column before the final synthesis of the [15O]water. This provided a practical way of removing tritium from the [15O]water.

Frequent expression of the variant CD30 in human malignant myeloid and lymphoid neoplasms.

We earlier identified a variant of CD30 (CD30v) that retains only the cytoplasmic region of the authentic CD30. This variant is expressed in alveolar macrophages. CD30v can activate the nuclear factor-kappaB (NF-kappaB) as CD30, and its overexpression in HL-60 induced a differentiated phenotype. To better understand the physiological and pathological functions of CD30v, expression of this variant was examined using a multiple approach to examine 238 samples of human malignant myeloid and lymphoid neoplasms. Screening by reverse transcriptase-polymerase chain reaction (RT-PCR) revealed expression of CD30v transcripts in 52 of 72, 7 of 11, 63 of 90, and 7 of 30 samples of acute myeloid leukemia (AML), myeloid blast crisis of myeloproliferative disorders (MBC), and lymphoproliferative disorders (LPDs) of B- and T-cell origin, respectively. CD30v expression was high in monocyte-oriented AMLs (FAB M4 and M5), B-cell chronic lymphocytic leukemia (B-CLL), and multiple myeloma (MM). Using the specific antibody HCD30C2, prepared using a peptide corresponding to the nine amino acids of the amino-terminal CD30v, expression of CD30v protein was detected in 10 of 25 and 2 of 10 AML and ALL samples, respectively. In AMLs, immunocytochemical detection of CD30v revealed the presence of loose clusters of CD30v-expressing cells dispersed amid a population of CD30v-negative blasts. Finally, the parallel expression of CD30v mRNA and protein, as evidenced by Northern and Western blotting, was confirmed in selected cases of AMLs and LPDs. A significant correlation was found between expressions of CD30v and CD30 ligand transcripts in AML and LPD (P = 0.02, odds ratio = 3.2). The association of CD30v with signal-transducing proteins, tumor necrosis factor receptor-associated factor (TRAF) 2, and TRAF5 was demonstrated by coimmunoprecipitation analysis, as was demonstrated for authentic CD30 protein. Expression of transcripts for TRAF1, TRAF2, TRAF3, and TRAF5, as demonstrated by RT-PCR, was noted in leukemic blasts that express CD30v. Collectively, frequent expression of CD30v along with TRAF proteins in human neoplastic cells of myeloid and lymphoid origin provide supportive evidence for biological and possible pathological functions of this protein in the growth and differentiation of a variety of myeloid and lymphoid cells.

Abnormalities of the p53 gene in juvenile myelomonocytic leukaemia.

Juvenile chronic myelomonocytic leukaemia (JMML) is a rare myeloproliferative disorder of childhood. Fewer than 30% of cases of JMML terminate in a blast crisis; however, its molecular mechanism is unknown. Since mutation and/or deletion of the p53 gene has been reported to be associated with disease progression in a wide variety of human cancers, including adult-type chronic myelogenous leukaemia, we studied the p53 gene in 20 patients with JMML (16 samples in chronic phase and seven at blast crisis). Exons 4-8 of the p53 gene, which cover all the hot spots of point mutations, were amplified by the polymerase chain reaction (PCR) method and subjected to mutation screening by single-strand conformation polymorphism analysis. No mobility shift of single-strand DNA of PCR products in polyacrylamide gel electrophoresis, indicating point mutations, was found in 19/20 patients. DNA of the remaining patient in the chronic phase failed to be amplified by PCR and Southern blot analysis with XbaI-digested genomic DNA revealed a gross rearrangement (presumed deletion) of the p53 gene. These data indicate that abnormalities of the p53 gene are rare in JMML and not responsible for acute transformation, but could be involved in the pathogenesis of some cases of JMML.

All-trans retinoic acid and hematopoietic growth factors regulating the growth and differentiation of blast progenitors in acute promyelocytic leukemia.

Although acute leukemia is generally thought to be characterized by maturation arrest, it has been shown that differentiation occurs in blast cells of acute myelogenous leukemia (AML) in vitro as well as in vivo, and that morphologically abnormal mature polymorphonuclear neutrophils (PMNs) often seen in patients with AML are possibly derived from spontaneously differentiating leukemic cells. Acute promyelocytic leukemia (APL) is an unique example in which these features of AML are evident in an almost complete form; administration of all-trans retinoic acid (ATRA) induces differentiation of neoplastic cells into mature neutrophils and successfully induce complete remission in most patients. However, PMNs appearing during ATRA treatment are morphologically abnormal, as indicated not only by the presence of Auer rods but also by neutrophil secondary-granule deficiency that is commonly seen in AML. Moreover, ATRA has heterogeneous effects on the growth of blast progenitors in APL in different patients, being inhibitory, stimulatory or ineffective, which might account in part for the leukemia relapse in patients treated with ATRA alone. Hematopoietic growth factors regulate the growth of blast progenitors in APL. Among them, granulocyte colony-stimulating factor (G-CSF) is unique in that it preferentially stimulates clonal growth, but not self-renewal, in many APL cases, and synergistically enhances the differentiation-inducing effect of ATRA when used in combination. Many other compounds also exert such synergistic effects with ATRA, for which a variety of mechanisms have been suggested. It is crucial to precisely elucidate the functions of these molecules governing the growth/differentiation balance of AML blast progenitors and the mechanisms underlying their deregulated differentiation program in order to achieve effective differentiation therapy for patients with AML, not restricted to APL.

Histopathologic findings of advanced neuroblastoma after intensive induction chemotherapy.

BACKGROUND: Histopathologic findings of advanced neuroblastoma after intensive induction chemotherapy have not been studied well. METHODS: In the present study, all of the surgical specimens from 19 patients who had advanced abdominal neuroblastoma and were pretreated intensively with the protocol of the Study Group of Japan were reviewed. The authors found that dissection of the contralateral lymph nodes is mandatory in advanced neuroblastoma when the goal is the complete dissection of the abdominal disease. Effects of chemotherapy were graded histologically according to the ratio of viable residual neuroblastoma tissue to total areas of the tumor, including neuroblastoma, ganglioneuroblastoma, ganglioneuroma, hemorrhage, necrosis and fibrosis, in five ranks from ( ) to (-). CONCLUSIONS: The newly introduced, highly cytotoxic regimen of the Japanese protocol, designated "A3," appears to be more effective histologically than the conventional regimen, designated "A1" or "new A1." Effects designated ( ) or (++) were prerequisites for survival in stage IV disease, but some stage III patients with the (+) effect survived.

Trends of survival in neuroblastoma and independent risk factors for survival at a single institution.

To assess the progress of survival in neuroblastoma which varies with many risk factors and to evaluate the influence of these factors on survival as independent risk factors. The study subjects were 159 neuroblastoma patients seen from 1965-1994 at the oldest and largest children's hospital in Japan. Trends of survival in three treatment eras-1965-81, 1982-86, 1987-94-were assessed by the Kaplan-Meier method for different sex, age at diagnosis, the clinical stage, the site of onset, and the histological type. Then the influence on survival of these factors as independent prognostic variables was evaluated by the Cox proportional hazards regression analysis. Age at diagnosis, the clinical stage, the site of onset, the histological type, and the treatment era were independent risk factors in the order of their influence on survival. Unfavorable survival outcomes were obtained for patients with age at diagnosis above 1 year, the clinical stage of VI by the Evans classification, adrenal onset, and neuroblastoma rather than ganglioneuroblastoma. Survival improved from the first to the second and from the second to the third treatment era. Improvement of survival in neuroblastoma took place during the past 3 decades. Age at diagnosis, the clinical stage, and the histological type have still remained overwhelming prognostic factors over the progress in treatment.

Neutrophil secondary-granule deficiency as a hallmark of all-trans retinoic acid-induced differentiation of acute promyelocytic leukemia cells.

Acute promyelocytic leukemia (APL) is a neoplasm with the unique chromosomal translocation t(15;17), which involves the retinoic acid receptor alpha gene. All-trans retinoic acid (ATRA) has been used for APL patients as a potent therapeutic agent to induce differentiation of leukemia cells. Although polymorphonuclear leukocytes (PMNs) appearing in the blood and bone marrow during ATRA treatment often possess Auer rods, indicating their neoplastic origin, other morphological abnormalities of PMNs have not been elucidated. We studied the morphological changes of APL cells during ATRA treatment at the ultrastructural level. Although most aberrant primary granules, including Auer rods, became morphologically normal in response to ATRA therapy and the nuclei showed chromatin condensation and lobulation, resulting in the emergence of PMNs, the lobulated nuclei often had nuclear filamentous connections and/or nuclear blebs, indicating some pathological process. Furthermore, PMNs, particularly early in ATRA treatment, lacked neutrophil secondary granules as did the PMNs appearing in a culture of APL cells incubated with ATRA, findings consistent with previously reported data that acute myeloid leukemia cell lines do not produce secondary granule proteins even after induction of differentiation towards mature neutrophils. The present data indicate that ATRA is incapable of inducing complete morphological maturation of APL cells and that secondary-granule deficiency may be a hallmark of aberrantly differentiated leukemic cells.