Insulinoma: A retrospective study analyzing the differences between benign and malignant tumors.

BACKGROUND/OBJECTIVES: Insulinoma is a rare pancreatic tumor and, usually, a benign disease but can be a malignant one and, sometimes, a highly aggressive disease. The aim of this study was to determine differences between benign and malignant tumors. METHODS: Retrospective study of 103 patients with insulinoma treated in a tertiary center. It was analyzed demographic, clinical, laboratory, localization and histologic analysis of tumor and follow up data of subjects in order to identify differences between individuals benign and malignant disease. RESULTS: Almost all patients (87%) had a benign tumor and survival rates of 100% following pancreatic tumor surgery. Those with malignant tumors (13%) have a poor prognosis, 77% insulinoma-related deaths over a period of 1-300 months after the diagnosis with a survival rate of 24% in five years. The following factors are associated with an increased risk of malignant disease: duration of symptoms < 24 months, fasting time for the occurrence of hypoglycemia < 8 h, blood plasma insulin concentration ≥ 28 µU/mL and C-peptide ≥ 4.0 ng/mL at the glycemic nadir and tumor size ≥ 2.5 cm. CONCLUSIONS: Our data help to base the literature about these tumors, reinforcing that although insulinoma is usually a single benign and surgically treated neoplasia, the malignant one is difficult to treat. We highlight the data that help predict a malignancy behavior of tumor and suggest a long follow up after diagnosis in these cases.

Effects of antidiabetes drugs on functional independence measure on a subacute rehabilitation ward for stroke patients.

It has been reported that the improvement of activities of daily living (ADL) by rehabilitation affects glycemic control. However, there are no reports about antidiabetes drugs as factors affecting the outcomes of rehabilitation. Therefore, we investigated the effects of antidiabetes drugs on functional independence measure (FIM) [total (T), motor (M), and cognition (C) items] in stroke patients with diabetes who were discharged from the subacute rehabilitation ward. We chose the frequently used antidiabetes drugs [sulfonylurea (SU), dipeptidyl peptidase-IV inhibitors (DPP-IVIs), and α-glycosidase inhibitors (α-GIs)] as the basis for categorizing the patients. We compared the patients' background features and laboratory data among the three groups. As a result, when SU was used in stroke patients with diabetes, it is difficult to obtain significant FIM-M gain, FIM-C gain, FIM-M efficiency, and FIM-C efficiency compared with of-GIs. As a reason for this, we hypothesize the possibility of the involvement of insulin resistance. Therefore, we consider that insulin resistance should be determined early and that it is important to reduce insulin resistance comprehensively by involving experts.

22q11.2 deletion carriers and schizophrenia-associated novel variants.

The penetrance of schizophrenia risk in carriers of the 22q11.2 deletion is high but incomplete, suggesting the possibility of additional genetic defects. We performed whole exome sequencing on two individuals with 22q11.2 deletion, one with schizophrenia and the other who was psychosis-free. The results revealed novel genetic variants related to neuronal function exclusively in the person with schizophrenia (frameshift: KAT8, APOH and SNX31; nonsense: EFCAB11 and CLVS2). This study paves the way towards a more complete understanding of variant dose and genetic architecture in schizophrenia.

Pharmacoeconomic analysis of DPP-4 inhibitors.

Dipeptidyl peptidase-4 (DPP-4) inhibitors and other incretin-related drugs have attracted attention as antidiabetic agents, but they are expensive. The Japanese government has adopted a policy of reducing healthcare costs, and medical institutions must provide medical care while considering economic efficiency. This study was a comparative survey of the usage, treatment effectiveness, and cost of DPP-4 inhibitors. The subjects were patients prescribed DPP-4 inhibitors (sitagliptin, vildagliptin, and alogliptin) at Gifu Municipal Hospital between February 2010 and August 2011. HbA1c: Japan Diabetes Society values (%) and concomitant antidiabetic agents were surveyed for 12 weeks after the start of DPP-4 inhibitors. A cost-effectiveness analysis showed that the cost required for a 0.1% decrease in HbA1c for 12 weeks was the lowest with vildagliptin (2,478 yen; decrease in HbA1c: 0.75% +/- 0.85%). In a cost analysis with a virtual cohort of 1000 patients, the number of patients who achieved the treatment target (HbA1c 6.5%) was estimated with respect to a virtual cohort created based on the HbA1c level (7.59 +/- 1.13%) at baseline of 307 patients, in cases assuming the use of each DPP-4 inhibitor. In addition, the incremental cost-effectiveness ratio (ICER) was obtained with sitagliptin 50 mg as the reference. The number of patients achieving the treatment target was the highest with vildagliptin 100 mg (413 of 1000 patients), and the estimated ICER of 28,359 yen was the lowest. Robustness was also confirmed with a sensitivity analysis. These results suggest that vildagliptin provides a superior cost-benefit.

p21 provides stage specific DNA damage control to preimplantation embryos.

The early stage embryogenesis of higher eukaryotes lacks some of the damage response pathways such as G1/S checkpoint, G2/M checkpoint and apoptosis. We examined here the damage response of preimplantation stage embryos after fertilization with 6 Gy irradiated sperm. Sperm-irradiated embryos developed normally for the first 2.5 days, but started to exhibit a developmental delay at day 3.5. p21 was activated in the delayed embryos, which carried numerous micronuclei owing to delayed chromosome instability. Apoptosis was observed predominantly in the inner cell mass of the day 4.0 embryos. Sperm-irradiated p21-/- embryos lacked the delay, but chromosome instability and apoptosis were more pronounced than the corresponding p21 wild-type embryos. We conclude from the result that damage responses come in a stage-specific manner during preimplantation stage development; p53-dependent S checkpoint at the zygote stage, p21-mediated cell cycle arrest at the morula/blastocyst stages and apoptosis after the blastocyst stage in the inner cell mass.

Suppression of replication fork progression in low-dose-specific p53-dependent S-phase DNA damage checkpoint.

The S-phase DNA damage checkpoint is activated by DNA damage to delay DNA synthesis allowing time to resolve the replication block. We previously discovered the p53-dependent S-phase DNA damage checkpoint in mouse zygotes fertilized with irradiated sperm. Here, we report that the same p53 dependency holds in mouse embryonic fibroblasts (MEFs) at low doses of irradiation. DNA synthesis in p53 wild-type (WT) MEFs was suppressed in a biphasic manner in which a sharp decrease below 2.5 Gy was followed by a more moderate decrease up to 10 Gy. In contrast, p53-/- MEFs exhibited radioresistant DNA synthesis below 2.5 Gy whereas the cells retained the moderate suppression above 5 Gy. DNA fiber analysis revealed that 1 Gy irradiation suppressed replication fork progression in p53 WT MEFs, but not in p53-/- MEFs. Proliferating cell nuclear antigen (PCNA), clamp loader of DNA polymerase, was phosphorylated in WT MEFs after 1 Gy irradiation and redistributed to form foci in the nuclei. In contrast, PCNA was not phosphorylated and dissociated from chromatin in 1 Gy-irradiated p53-/- MEFs. These results demonstrate that the novel low-dose-specific p53-dependent S-phase DNA damage checkpoint is likely to regulate the replication fork movement through phosphorylation of PCNA.

Chromosome aberrations do not persist in the lymphocytes or bone marrow cells of mice irradiated in utero or soon after birth.

Mice were exposed at various ages to 1 Gy or 2 Gy of X rays, and translocation frequencies in peripheral blood T cells, spleen cells, and bone marrow cells were determined with FISH painting of chromosomes 1 and 3 when the animals were 20 weeks old. It was found that the mean translocation frequencies were very low (< or =0.8%) in mice exposed in the fetal or early postnatal stages. However, with the increase in animal age at the time of irradiation, the frequency observed at 20 weeks old became progressively higher then reached a plateau (about 5%) when mice were irradiated when > or =6 weeks old. A major role of p53 (Trp53)-dependent apoptosis for elimination of aberrant cells was not suggested because irradiated fetuses, regardless of the p53 gene status, showed low translocation frequencies (1.8% in p53(-/-) mice and 1.4% in p53(+/-) mice) compared to the frequency in the p53(-/-) mother (7.4%). In contrast, various types of aberrations were seen in spleen and liver cells when neonates were examined shortly after irradiation, similar to what was observed in bone marrow cells after irradiation in adults. We interpreted the results as indicating that fetal cells are generally sensitive to induction of chromosome aberrations but that the aberrant cells do not persist because fetal stem cells tend to be free of aberrations and their progeny replace the pre-existing cell populations during the postnatal growth of the animals.

Polyunsaturated fatty acid deficiency during neurodevelopment in mice models the prodromal state of schizophrenia through epigenetic changes in nuclear receptor genes.

The risk of schizophrenia is increased in offspring whose mothers experience malnutrition during pregnancy. Polyunsaturated fatty acids (PUFAs) are dietary components that are crucial for the structural and functional integrity of neural cells, and PUFA deficiency has been shown to be a risk factor for schizophrenia. Here, we show that gestational and early postnatal dietary deprivation of two PUFAs-arachidonic acid (AA) and docosahexaenoic acid (DHA)-elicited schizophrenia-like phenotypes in mouse offspring at adulthood. In the PUFA-deprived mouse group, we observed lower motivation and higher sensitivity to a hallucinogenic drug resembling the prodromal symptoms in schizophrenia. Furthermore, a working-memory task-evoked hyper-neuronal activity in the medial prefrontal cortex was also observed, along with the downregulation of genes in the prefrontal cortex involved in oligodendrocyte integrity and the gamma-aminobutyric acid (GABA)-ergic system. Regulation of these genes was mediated by the nuclear receptor genes Rxr and Ppar, whose promoters were hyper-methylated by the deprivation of dietary AA and DHA. In addition, the RXR agonist bexarotene upregulated oligodendrocyte- and GABA-related gene expression and suppressed the sensitivity of mice to the hallucinogenic drug. Notably, the expression of these nuclear receptor genes were also downregulated in hair-follicle cells from schizophrenia patients. These results suggest that PUFA deficiency during the early neurodevelopmental period in mice could model the prodromal state of schizophrenia through changes in the epigenetic regulation of nuclear receptor genes.

Involvement of M(2) muscarinic receptors in relaxant response of circular muscle of mouse gastric antrum.

Our previous study showed that atropine significantly inhibited the sustained relaxation induced by electrical field stimulation (EFS) in the circular muscle strips prepared from the mouse antrum, and that pituitary adenylate cyclase activating peptide (PACAP) partially mediated the sustained relaxation. The muscarinic receptor subtype associated with the sustained relaxation was examined in the present study by using each muscarinic receptor subtype of knockout (KO) mice. EFS-induced sustained relaxation in the antrum prepared from M(2) receptor KO mice was significantly less than that of wild-type mice. Atropine failed to inhibit this relaxation. On the other hand, similar sustained relaxation and inhibitory effects of atropine to those of wild-type mice were observed in M(1), M(3) and M(4) receptor KO mice. Exogenously added PACAP-27 relaxed the antral strips of wild-type and M(2) receptor KO mice to a similar extent. Immunohistochemical study revealed that M(2) receptor immunoreactivity was localized with PACAP-immunoreactivity in enteric neurons within the antrum of wild-type mice. In contrast, atropine did not affect the EFS-induced sustained relaxation in the gastric fundus. These results suggest that M(2) receptors modulate the sustained relaxation, probably through the regulation of PACAP release, in the mouse antrum.

Purification and characterization of the platelet-aggregating sialoglycoprotein gp44 expressed by highly metastatic variant cells of mouse colon adenocarcinoma 26.

A platelet-aggregating sialoglycoprotein with a molecular weight of 44,000 (gp44) was immunochemically purified from highly metastatic mouse adenocarcinoma cells. The rat monoclonal antibody (mAb) 8F11 used in the purification procedure has been generated previously against NL-17 cells derived from the mouse colon 26 cell line, mAb 8F11 inhibits NL-17 cells from inducing platelet aggregation and suppresses their experimental metastasis to the lung. The purified gp44 induced mouse platelet aggregation in a dose-dependent manner without any plasma component. This aggregation was completely inhibited by mAb 8F11. The gp44 was partially characterized by sequential enzymatic hydrolysis of carbohydrates and was found to be O-glycans enriched. When gp44 was sequentially treated with N-glycanase and neuraminidase, it lost platelet aggregation activity. Further treatment with O-glycanase resulted in a loss of the reactivity to mAb 8F11. These results suggest that sialylated carbohydrate chains of gp44 are involved in the induction of platelet aggregation and may play an important role in the colonization of NL-17 cells in the lung.

Transferrin facilitates the formation of DNA double-strand breaks via transferrin receptor 1: the possible involvement of transferrin in carcinogenesis of high-grade serous ovarian cancer.

Fallopian tubal epithelium is a candidate for the origin of high-grade serous ovarian cancer. Transferrin-containing follicular fluid and/or retrograde menstrual blood are possible risk factors for carcinogenesis. Accumulation of DNA double-strand breaks (DNA-DSBs) in the fallopian tubal epithelium is considered to play an important role in the development of cancer. However, the mechanisms by which DNA-DSBs accumulate have not yet been fully elucidated. The hydroxyl radical, which is produced in a Fenton reaction catalyzed by an iron ion, serves as a potent DNA-DSB-inducing molecule, raising the potential of an iron ion transporter of transferrin in the formation of DNA-DSBs. We studied the potential involvement of transferrin in DNA damage and the development of ovarian cancer. Treatment with transferrin facilitated the formation of histone 2AX phosphorylated at Serine 139 (γH2AX), which is known as a DNA-DSB marker, in human fallopian tube secretory epithelial cells and A2780 ovarian cancer cells. Knockdown of transferrin receptor 1 (TfR1), but not transferrin receptor 2, suppressed the transferrin uptake and consequent formation of γH2AX. As hydroxyl radicals in reactive oxygen species (ROS) are involved in DNA-DSBs, the formation of ROS was determined. Treatment with TfR1-specific small interference RNAs significantly diminished transferrin-induced formation of ROS. Moreover, TfR1-dependent uptake of transferrin was revealed to augment the formation of DNA-DSBs in the presence of hydrogen peroxide, which served as a substrate for the Fenton reaction. An ex vivo study with murine fallopian tubes further demonstrated that transferrin treatment introduced DNA-DSBs in the fallopian tubal epithelium. Collectively, these data suggested that the transferrin-TfR1 axis accounts for the induction of DNA-DSBs that potentially lead to DNA damage/genome instability. These findings also suggested that exposure to transferrin initiates and promotes the development of ovarian cancer by aiding the accumulation of DNA-DSBs in the fallopian tubal epithelium.

Ceramide limits phosphatidylinositol-3-kinase C2ß-controlled cell motility in ovarian cancer: potential of ceramide as a metastasis-suppressor lipid.

Targeting cell motility, which is required for dissemination and metastasis, has therapeutic potential for ovarian cancer metastasis, and regulatory mechanisms of cell motility need to be uncovered for developing novel therapeutics. Invasive ovarian cancer cells spontaneously formed protrusions, such as lamellipodia, which are required for generating locomotive force in cell motility. Short interfering RNA screening identified class II phosphatidylinositol 3-kinase C2ß (PI3KC2ß) as the predominant isoform of PI3K involved in lamellipodia formation of ovarian cancer cells. The bioactive sphingolipid ceramide has emerged as an antitumorigenic lipid, and treatment with short-chain C6-ceramide decreased the number of ovarian cancer cells with PI3KC2ß-driven lamellipodia. Pharmacological analysis demonstrated that long-chain ceramide regenerated from C6-ceramide through the salvage/recycling pathway, at least in part, mediated the action of C6-ceramide. Mechanistically, ceramide was revealed to interact with the PIK-catalytic domain of PI3KC2ß and affect its compartmentalization, thereby suppressing PI3KC2ß activation and its driven cell motility. Ceramide treatment also suppressed cell motility promoted by epithelial growth factor, which is a prometastatic factor. To examine the role of ceramide in ovarian cancer metastasis, ceramide liposomes were employed and confirmed to suppress cell motility in vitro. Ceramide liposomes had an inhibitory effect on peritoneal metastasis in a murine xenograft model of human ovarian cancer. Metastasis of PI3KC2ß knocked-down cells was insensitive to treatment with ceramide liposomes, suggesting specific involvement of ceramide interaction with PI3KC2ß in metastasis suppression. Our study identified ceramide as a bioactive lipid that limits PI3KC2ß-governed cell motility, and ceramide is proposed to serve as a metastasis-suppressor lipid in ovarian cancer. These findings could be translated into developing ceramide-based therapy for metastatic diseases.

Analysis of induced pluripotent stem cells carrying 22q11.2 deletion.

Given the complexity and heterogeneity of the genomic architecture underlying schizophrenia, molecular analyses of these patients with defined and large effect-size genomic defects could provide valuable clues. We established human-induced pluripotent stem cells from two schizophrenia patients with the 22q11.2 deletion (two cell lines from each subject, total of four cell lines) and three controls (total of four cell lines). Neurosphere size, neural differentiation efficiency, neurite outgrowth, cellular migration and the neurogenic-to-gliogenic competence ratio were significantly reduced in patient-derived cells. As an underlying mechanism, we focused on the role of DGCR8, a key gene for microRNA (miRNA) processing and mapped in the deleted region. In mice, Dgcr8 hetero-knockout is known to show a similar phenotype of reduced neurosphere size (Ouchi et al., 2013). The miRNA profiling detected reduced expression levels of miRNAs belonging to miR-17/92 cluster and miR-106a/b in the patient-derived neurospheres. Those miRNAs are reported to target p38α, and conformingly the levels of p38α were upregulated in the patient-derived cells. p38α is known to drive gliogenic differentiation. The inhibition of p38 activity by SB203580 in patient-derived neurospheres partially restored neurogenic competence. Furthermore, we detected elevated expression of GFAP, a gliogenic (astrocyte) marker, in postmortem brains from schizophrenia patients without the 22q11.2 deletion, whereas inflammation markers (IL1B and IL6) remained unchanged. In contrast, a neuronal marker, MAP2 expressions were decreased in schizophrenia brains. These results suggest that a dysregulated balance of neurogenic-to-gliogenic competence may underlie neurodevelopmental disorders such as schizophrenia.

Genomic organization and chromosome localization of the newly identified human heparanase gene.

Heparanase (HPSE), which we have recently isolated, is an endo-beta-D-glucuronidase capable of cleaving heparan sulfate and has been implicated in inflammation and tumor angiogenesis and metastasis. In this report, the genomic organization and chromosome localization of the human heparanase gene is described. Polymerase chain reaction, subcloning and DNA sequencing analysis of a bacterial artificial chromosome (BAC) clone revealed that the 3.7 kb human heparanase cDNA is spread over about 50 kb and contains 14 exons and 13 introns. The heparanase gene is expressed as two mRNA species containing the same open reading frame, HPSE 1a (5 kb) (GenBank Data Library under accession number: AF155510); and HPSE 1b (1.7 kb) (GenBank Data Library under accession number: AF144325), generated by alternative splicing. The HPSE 1a-form contains all 14 exons, whereas in the HPSE 1b-form the first and fourteenth exons (5'- and 3'-untranslated region) have been spliced out. All splice sites conform to the GT-AG rule, except for the splice donor site of intron 13 (which is GA instead of GT), and the splice acceptor of intron 13 (which is GG instead of AG). Fluorescence in situ hybridization and radiation hybrid mapping suggest that the heparanase gene is located on human chromosome 4q22. This report regarding the structure of the human heparanase gene will aid in understanding the genetic contribution of this gene to normal physiology as well as to disease states. A possible involvement of heparanase in neuronal degeneration is discussed.

Inhibition of tumor invasion and metastasis by calcium spirulan (Ca-SP), a novel sulfated polysaccharide derived from a blue-green alga, Spirulina platensis.

We have investigated the effect of calcium spirulan (Ca-SP) isolated from a blue-green alga, Spirulina platensis, which is a sulfated polysaccharide chelating calcium and mainly composed of rhamnose, on invasion of B16-BL6 melanoma, Colon 26 M3.1 carcinoma and HT-1080 fibrosarcoma cells through reconstituted basement membrane (Matrigel). Ca-SP significantly inhibited the invasion of these tumor cells through Matrigel/fibronectin-coated filters. Ca-SP also inhibited the haptotactic migration of tumor cells to laminin, but it had no effect on that to fibronectin. Ca-SP prevented the adhesion of B16-BL6 cells to Matrigel and laminin substrates but did not affect the adhesion to fibronectin. The pretreatment of tumor cells with Ca-SP inhibited the adhesion to laminin, while the pretreatment of laminin substrates did not. Ca-SP had no effect on the production and activation of type IV collagenase in gelatin zymography. In contrast, Ca-SP significantly inhibited degradation of heparan sulfate by purified heparanase. The experimental lung metastasis was significantly reduced by co-injection of B16-BL6 cells with Ca-SP. Seven intermittent i.v. injections of 100 microg of Ca-SP caused a marked decrease of lung tumor colonization of B16-BL6 cells in a spontaneous lung metastasis model. These results suggest that Ca-SP, a novel sulfated polysaccharide, could reduce the lung metastasis of B16-BL6 melanoma cells, by inhibiting the tumor invasion of basement membrane probably through the prevention of the adhesion and migration of tumor cells to laminin substrate and of the heparanase activity.

Ataxia-telangiectasia without immunodeficiency: novel point mutations within and adjacent to the phosphatidylinositol 3-kinase-like domain.

Ataxia-telangiectasia (AT) is an autosomal recessive disorder characterized by progressive ataxia, telangiectasia, sinopulmonary infections, hypersensitivity to ionizing radiation, and combined immunodeficiency. Recently, the AT gene (ATM) was cloned and shown to be mutated in AT patients. In this report, mutation analysis of ATM was performed in a 24-year-old AT patient without immunodeficiency. ATM amplified with reverse transcriptase-polymerase chain reaction (RT-PCR) was screened with a ribonuclease (RNase) cleavage assay and auto-sequenced. This patient, a compound heterozygote, showed two mutations in ATM: one missense mutation leading to a Leu2656Pro substitution and the other to the truncation at codon 3047 (Arg-->ter). The latter mutation is within the phosphatidylinositol 3-kinase (PI 3-kinase)-like domain and the former is outside but close to the domain. The particular phenotype in our patient, no immunodeficiency, suggests incomplete functional loss of ATM protein. The clinical spectrum of AT caused by ATM mutations may be broader than previously thought. Further analysis of patients with similar phenotypes will make the relation between ATM genotype and phenotype clear.

Mutual alignment errors due to the variation of wave-front aberrations in a free-space laser communication link.

Optical devices in free-space laser communication systems are affected by their environment, particularly in relation to the effects of temperature while in orbit. The mutual alignment error between the transmitted and received optical axes is caused by deformation of the optics due to temperature variation in spite of the common optics used for transmission and reception of the optical beams. When a Gaussian beam wave for transmission is aligned at the center of a received plane wave, 3rd-order Coma aberrations have the most influence on the mutual alignment error, which is an inevitable open pointing error under only the Tip/Tilt tracking control. As an example, a mutual alignment error of less than 0.2 microrad is predicted for a laser communication terminal in orbit using the results from space chamber thermal vacuum tests. The relative power penalty due to aberration is estimated to be about 0.4 dB. The results will mitigate surface quality in an optical antenna and contribute to the design of free-space laser communication systems.

Wegener's granulomatosis responding to antituberculous drugs.

We present a case of Wegener's granulomatosis (WG) that responded to antituberculous drugs. A 44-year-old woman with multiple nodules on chest radiograph received a diagnosis of pulmonary tuberculosis because open-lung biopsy specimens showed caseous granulomas. Her chest shadows underwent repeated resolution after the start of antituberculous treatment, and relapse after the cessation of the drugs. Antineutrophil cytoplasmic antibody was positive (14 enzyme-linked immunosorbent assay units), and the second lung biopsy specimens showed necrotizing granulomas and vasculitis without pathogenic organisms. Thus, the patient received a diagnosis of WG and was successfully treated with trimethoprim/sulfamethoxazole 10 years after her initial evaluation. Antituberculous drugs were effective in this case of WG.

A clinical study of idiopathic eosinophilic pneumonia.

To better characterize idiopathic eosinophilic pneumonia (IEP), we studied the clinical and laboratory features of 27 patients. Patients with IEP could be divided into those with chronic eosinophilic pneumonia (CEP) (n = 14) and acute eosinophilic pneumonia (AEP) (n = 13). CEP was characterized by (1) multiple and dense areas of consolidation on chest radiographs and computed tomographic (CT) scans, (2) persistent symptoms, (3) a requirement for steroid therapy, and (4) possible relapses. On the other hand, AEP was characterized by (1) diffuse ground-glass and micronodular infiltrates on radiographs and CT scans (in mild cases, the lesions were sparse or localized), (2) acute onset with high fever, (3) spontaneous improvement, and (4) no relapse. In addition, peripheral blood eosinophil count was significantly higher in patients with CEP than in patients with AEP at the first examination. However, the eosinophil fraction also became markedly elevated during the subsequent courses of AEP. Analysis of bronchoalveolar lavage fluid revealed that the percentage of eosinophils was higher in patients with CEP than that in patients with AEP, whereas the percentage of lymphocytes was significantly greater in patients with AEP than patients with CEP. It was also noted that 75 percent of patients with CEP and 82 percent of patients with AEP had allergic diathesis, suggesting that both conditions are likely to occur in atopic individuals.

Determination of manganese in tissues by neutron activation analysis using an antimony pentoxide column.

A rapid and accurate method is presented for the determination of manganese in biological samples, using neutron activation analysis. Biological samples were irradiated at 5000 kW for 30 min. The samples were ashed on a hot plate with 14 mol/l HNO3 and 6 mol/l HClO4, and resolved in 1 mol/l HClO4. 24Na and 24K were removed by passing each sample through in antimony pentoxide column. 54Mn was added as a tracer to calculate the ratio of manganese recovered by the separation procedure. Recovery was over 90%. This method was applied in order to determine manganese in various tissues. In the cervical spinal cord of the controls, the mean manganese concentrations in the anterior horn, the lateral and the posterior columns were 1.14, 1.06 and 0.90 ng/mg of dried tissue, respectively. In two cases if amyotrophic lateral sclerosis the manganese concentrations in the cervical spinal cord were elevated, particularly in the anterior horn and the lateral column.