Activity-dependent survival and enhanced turnover of calcium in cultured rat cerebellar granule neurons

Neurons survive when their activity is maintained. An influential hypothesis on the cellular mechanism underlying this phenomenon is that there is an appropriate range of intracellular Ca²⁺ concentration ([Ca²⁺]i) for survival. The rat cerebellar granule neuron in culture serves as the most often used model system for the analysis of activity-dependent survival, since it does not survive unless an excitant (KCl or glutamate) is added to the culture medium. Against the above-mentioned hypothesis, we found in our previous examination no difference between steady-state [Ca²⁺]i in granule neurons cultured under high KCl (i.e., survival) and low KCl (i.e., death) conditions. In this report, we present the quantitative background of unchanged [Ca²⁺]i between the two culture conditions. Influx of Ca²⁺ due predominantly to L-type voltage-dependent calcium channels was higher in high KCl cultures than in low KCl cultures. At the same time, efflux of Ca²⁺ due to the activity of Ca²⁺/Na⁺ antiport was also higher in high KCl cultures. Additionally, we found that the endocytotic activity was greater in high KCl cultures than in low KCl cultures, as monitored by the rate of uptake of horseradish peroxidase added to medium. Since the uptake was blocked by an internal Ca²⁺ chelator, the increased endocytotic activity in high KCl cultures might be a consequence of the enhanced Ca²⁺ turnover.     

Development of new immunoradiometric assay for CA 125 antigen using two monoclonal antibodies produced by immunizing lung cancer cells

CA 125 is an antigen associated with non-mucinous epithelial ovarian cancer, which is defined by OC 125 antibody developed by immunizing ovarian cancer cells. We have produced two monoclonal antibodies, 130-22 and 145-9, by using the human lung adenocarcinoma cell line PC-9. Both 130-22 and 145-9 antibodies recognized CA 125 antigen. However, the binding sites seemed to be separate from those of OC 125. Testing by 9 immunoradiometric assays (IRMA), using different combinations of the 3 monoclonal antibodies 130-22, 145-9 and OC 125 demonstrated that the best standard curve for detecting CA 125 could be obtained by a "simultaneous sandwich" assay based on a mixture of 125I-labeled OC 125 and 130-22 or 145-9 coated beads. One-step IRMA, using 130-22 as a tracer and 145-9 as an immunoadsorbent, also showed good reproducibility and sensitivity for measuring CA 125. Antigens were detectable in the culture supernatants of PC-9 cells and 5 of 6 ovarian cancer and endometrial adenocarcinoma cells. These results indicate that one-step IRMA using 130-22 and 145-9 is useful for detecting CA 125 antigen.     

Growth and Production of Steroid Saponin in Solanum aculeatissimum During One Vegetation Period

SOLANUM ACULEATISSIMUM (Solanaceae) plants, which were cultivated in the open field, were harvested once a month from June 1985 to January 1986. The growth matter and content of steroid saponins, aculeatiside A and B which are precursors of steroid hormones in root, were then measured in order to examine their seasonal variations during one vegetation period. The aerial parts of the plants died of chill and frost in January. In November the dry weight of root reached the maximum, and the content of steroid saponins was more than 10% and showed a maximum. The yield of steroid saponins per plant was greatest in November. These results suggest that November is the appropriate time for harvest, and that S. ACULEATISSIMUM can be more useful than the DIOSCOREA species which is now used as a source plant of steroid hormones.     

A cysteine-activated protease isolated from Todarodes pacificus squid degrades collagen below its denaturation temperature

Collagen purified from the mantle muscle of the Japanese common squid, Todarodes pacificus, showed autodegradation during incubation under acidic conditions at 25 degrees C, without the addition of exogenous enzymes. This suggests that the collagenolytic proteases bind to collagen tightly through the steps of collagen preparation. Collagenolytic activity also was detected in a crude extract of mantle muscle, and leupeptin and E-64 were observed to inhibit collagenolytic activity within the collagen fraction and muscle extract. We purified these collagenolytic cysteine proteases by leupeptin column chromatography and cellulose acetate membrane electrophoresis. Optimal enzymatic activity was observed at pH 3.5, and collagenolytic activity was completely suppressed at neutral or alkaline pH. The purified enzymes were 28 kDa and 25 kDa in size, and both had gelatinolytic activity, as detected by gelatin zymography, and cut the specific site of denatured collagen alpha chain. The purified enzymes degraded squid collagen at 25 degrees C, which is 2.5 degrees lower than the temperature at which squid collagen normally denatures; however, the proteases were ineffective at 20 degrees C. Interestingly, the isolated proteases were capable of digesting both squid and bovine gelatin. In this article, we describe collagenolytic cysteine proteases that bind to the collagen of Todarodes pacificus, thereby digesting it by attacking microunfolding regions generated by incubation 2-3 degrees C below the denaturation temperature.     

Distant metastases in differentiated thyroid carcinomas: a clinical and pathologic study

Of 514 patients with differentiated thyroid carcinoma treated between 1970 and 1987, 34 (6.6%) had distant metastases. Twelve patients died of their distant metastases; eight of these patients died within 5 years from the time of initial diagnosis. Death from cancer was most frequent in the seventh decade. The metastases were most often found in the lungs and bones. In the fatal group, pleural, brain, and pericardial metastases were noted. Local recurrences were found only in 24% of these 34 patients. Histologic types of primary thyroid tumors and metastatic tumors were reexamined and classified using our criteria, which were mainly based on the World Health Organization nomenclature and currently obtained pathologic observations of thyroid tumors. In 31 thyroid tumors, the surgical specimens were available for review. Twenty-four tumors were papillary and seven were follicular. Of the 24 papillary carcinomas, nine were follicular, eight were well-differentiated, and seven were trabecular. On the other hand, the seven follicular carcinomas consisted of four well-differentiated, two solid, and one oxyphilic. The majority of the thyroid tumors showed an extrathyroidal extension; however, two were intrathyroidal carcinomas and two were encapsulated carcinomas, larger than 5 cm in diameter. Distant metastases were confirmed morphologically in 18 patients (11 by surgical or biopsy material, five by autopsy, and two by cytology). The histologic types of metastatic tumors were consistent with those of primary thyroid tumors. Diagnostic 131I uptake was examined in 32 patients and absorption of diagnostic 131I in metastatic tumors was demonstrated in 21 patients. The 10-year survival rate of patients with 131I accumulating metastases (70%) was significantly better than that of patients with metastases lacking such uptake (40%). Immunoreactivity for thyroglobulin in metastatic tumors was correlated with the 131I absorption. This finding indicated that immunostaining of thyroglobulin in metastatic tumors might be useful in the prediction of the effectiveness of 131I therapy.     

Neonate blood IgE levels on filter paper as indicators of atopic disease

Measurements of IgE levels in the blood of neonates were investigated using filter paper for blood collection in mass screening of congenital metabolic disorders. Time-resolved fluoroimmunometric assay system for the measurement of filter paper blood IgE levels was also studied. In an analysis of the present results, IgE values of at least 0.015U/ml, the measurement limit, were considered as high. High IgE levels in filter paper blood were seen in 28 (7.2%) of the 389 cases. When the relation with serum IgE levels at 18 months of age was investigated in 134 of 389 subjects, high serum IgE levels were also found in about 86.7% of the subjects with high IgE levels in filter paper blood. In addition, when the relation between family history of atopic disease and presence of atopic disease in the first 18 months of age was investigated in 203 of the 389 subjects, about 90% of the subjects with a family history of atopic disease and high IgE levels in filter paper blood developed atopic disease. Since filter paper blood is routinely collected in Japan, IgE levels in this blood should be widely checked for the prediction of onset of atopic disease in infants.     

Duct-Islet Cell Tumor of the Pancreas. A Case Report with Immunohistochemical and Electron Microscopic Findings

A case of pancreatic tumor with features of both duct and islet cell components was found incidentally at autopsy in a 76 year old male who had died of intrahepatic cholan-giocarcinoma. The tumor, measuring about l.0cm in diameter, was located in the pancreatic tail. The tumor was composed of two distinct cell populations, islet cells and duct cells. Immunocytochemically, nearly all of the former cells were positive for insulin but negative for cytokeratin, carcinoembryonic antigen (CEA) and mucin, while the latter were positive for cytokeratin, CEA and mucin but negative for insulin. Additionally, a majority of the tumor cells that had formed islet-like structures were positive for neuron specific enolase (NSE), whereas NSE-positive cells were found only rarely in duct components. Electron microscopy confirmed the presence of two cell populations. Simultaneous occurrence of duct and islet cell components in a single pancreatic tumor indicates an intimate histogenetic relationship between pancreatic endocrine and duct cells. Acta Pathol Jpn 39: 328 335, 1989.     

Usefulness of body surface mapping to differentiate patients with Brugada syndrome from patients with asymptomatic Brugada syndrome

We attempted to determine the usefulness of body surface mapping (BSM) for differentiating patients with Brugada syndrome (BS) from patients with asymptomatic Brugada syndrome (ABS). Electrocardiograms (ECG) and BSM were recorded in 7 patients with BS and 35 patients with ABS. Following the administration of Ic antiarrhythmic drugs, BSM was recorded in 5 patients with BS and 16 patients with ABS. The maximum amplitudes at J0, J20, J40 and J60 were compared between the 2 groups, as were 3-dimensional maps. The maximum amplitudes at J0, J20 and J60 under control conditions were larger in patients with BS than in patients with ABS (P < 0.05). A three-dimensional map of the ST segments under control conditions in patients with BS showed a higher peak of ST elevation in the median precordium compared to that for patients with ABS. Increases in ST elevation at J20, J40 and J60 following drug administration were greater in patients with BS than in patients with ABS (P < 0.05). Evaluation of the change in amplitude of the ST segment at E5 caused by Ic drug administration was also useful for differentiating between the 2 groups. In conclusion, BSM was useful for differentiating patients with BS from those with ABS.     

Immunochemical analysis of two peak M proteinemia derived from structural differences of IgG Fc region

We found M-proteins with two peaks by agarose electrophoresis in the serum of a myeloma patient. The M-proteins were identified as both IgG 1-kappa type, and classified as IgG-F (fast mobility) and IgG-S (slow mobility). 1) The possibility that the two M-proteins were derived from the post translational differences of sugar moieties of the same IgG molecule was unlikely, because no migration changes were observed in IgG-F and IgG-S after the treatment with 4 different sugar enzymes. 2) Fab fractions of IgG-F and IgG-S were analyzed. After papain or pepsin digestion, western blotting with anti-Fab antiserum revealed that the Fab fraction of IgG-F and IgG-S had identical mobility by agarose electrophoresis. However the Fc fractions of IgG-F and IgG-S analyzed by the same procedures with anti-Fe antiserum, were different. 3) Anti-idiotype antiserum prepared in rabbits against IgG-S, or -F, and absorbed by normal IgG and normal human serum showed a fused precipitin line with IgG-F and IgG-S. These findings suggest that two M-proteins with both IgG 1 and kappa type, have the same VH and VL regions but have different constant regions of heavy chain. Since one copy of IgG 1 constant gene is found in each human haploid gene. It is speculated that the switching of the rearranged VDJ gene to constant region gene occurred not only between cis chromosome but also between trans chromosome.     

AN AUTOPSY CASE OF PRIMARY ANGIOSARCOMA OF THE PERICARDIUM MIMICKING MALIGNANT MESOTHELIOMA

The pathology of a rare case of primary diffuse angiosarcoma of the pericardium is reported. Grossly, the heart was entirely encased by the pericardial tumor, and the myocardium was only superficially invaded by the tumor. The tumor tissue extended directly to the mediastinum, where the great vessels were embedded in the tumor. A few minute distant metastases were found only in the bilateral lungs and pulmonary hilar lymph nodes. Microscopically, the tumor tissue was composed of malignant cells forming vascular channels admixed with solid areas. Histo- and immunohistochemically, no mesothelial characteristics were evident. Factor VHI-related antigen and Ulex'europaeus I lectin were positive, implying that the tumor was of vascular origin. Grossly, and in part microscopically, this case resembled malignant diffuse mesothelioma, indicating that pericardial angiosarcoma may sometimes mimick malignant mesothelioma. ACTA PATHOL JPN 38: 1345-1351, 1988.