Immunohistochemical demonstration of DNA polymerase alpha in human brain-tumor cells

The proliferative capacity of brain-tumor cells was analyzed in vitro and in situ using monoclonal antibody (MAb) against deoxyribonucleic acid (DNA) polymerase alpha. For the in vitro studies, two cultured human glioma cell lines were investigated using MAb against DNA polymerase alpha, the MAb Ki-67, a serum against proliferating cell nuclear antigen (PCNA/cyclin), bromodeoxyuridine (BUdR), and an anti-BUdR MAb. During exponential growth of the cells, the percentage of polymerase alpha-positive cells (the "polymerase alpha score") ranged from 72.0% to 77.1%, the Ki-67-positive cells (the "Ki-67 score") ranged from 43.4% to 59.4%, the PCNA/cyclin-positive cells from 30.9% to 41.4%, and the BUdR labeling index from 28.6% to 39.3%. For the in situ studies, tissue from 60 human brain tumors and from two normal human brains was investigated and the polymerase alpha scores and Ki-67 scores were compared. In normal brain tissue, no immunostaining was found by either method. In brain tumors, both the polymerase alpha scores and the Ki-67 scores correlated with the histological grade of malignancy. Polymerase alpha scores were generally higher than Ki-67 scores in the same specimen, especially in malignant brain tumors. These findings suggest that immunostaining of DNA polymerase alpha is a convenient and important new method by which to estimate the cellular proliferation rate of brain tumors. Polymerase alpha scores may be closer to the growth fraction of the individual tumor than the MAb Ki-67 or other scores.     

Oral senile amyloidosis in senescence accelerated mouse (SAM)

Oral senile amyloidosis in senescence accelerated mouse (SAM) was examined for two SAM sublines (P/2/Iw and R/1/Iw) and for various ages by light microscopy, immunohistochemistry, and electron microscopy. The amyloid deposition, identified by green birefrigence following Congo red stain, was observed only in P/2/Iw. In P/2/Iw, no amyloid deposition was found at age 6 months; however, frequency and extent of such deposits increased with advancing age. Distribution of amyloid deposition was as follows: along papillary layers of mucous epithelium in the tongue, the gingiva, the palate, and the buccal mucosa; foci in connective tissues; along vessels, muscles, and minor salivary glands. Immunohistochemically, the amyloid deposition was positive with anti-ASSAM serum being raised against a unique amyloid protein ASSAM which probably induced "senile systemic amyloidosis". P/2/Iw is useful as an animal model of oral senile amyloidosis.     

Effect of irradiation on transforming growth factor-β secretion by malignant glioma cells

Glioblastoma cells secrete transforming growth factor- (TGF-), whichhas a variety of immunosuppressive properties. We investigatedthe effect of irradiation TGF- secretion by malignantglioma cells. Three malignant glioma cell lines (T98G,A172, KG-1-C) were cultured and irradiated using 10and 50 Gy Linac radiation. After further culturefor 36 hours in serum-free culture medium, thesupernatants were collected. The TGF- activity in theculture supernatants was determined using a specific bioassay.The levels of the active form and totalTGF- in the supernatants from irradiated malignant gliomacells decreased compared to those from un-irradiated cells.However, since irradiation inhibited the growth of tumorcells, the amount of TGF- secretion per cellin irradiated cells tended to increase after irradiation.These results suggest that malignant glioma cells canstill secrete TGF- and activate latent TGF- evenafter large dose irradiation, despite the inhibition oftumor growth.     

Inhibition of Ca2+ channel current by μ- and κ-opioid receptors coexpressed in Xenopus oocytes: desensitization dependence on Ca2+ channel α1 subunits

1. Desensitization of μ- and κ-opioid receptor-mediated inhibition of voltage-dependent Ca²⁺ channels was studied in a Xenopus oocyte translation system.
2. In the oocytes coexpressing κ-opioid receptors with N- or Q-type Ca²⁺ channel α₁ and β subunits, the κ-agonist, U50488H, inhibited both neuronal Ca²⁺ channel current responses in a pertussis toxin-sensitive manner and the inhibition was reduced by prolonged agonist exposure.
3. More than 10 min was required to halve the inhibition of Q-type channels by the κ-agonist. However, the half-life for the inhibition of N-type channels was only 6±1 min. In addition, in the oocytes coexpressing μ-opioid receptors with N-type or Q-type channels, the uncoupling rate of the μ-receptor-mediated inhibition of N-channels was also faster than that of Q-type channels.
4. In the oocytes coexpressing both μ- and κ-receptors with N-type channels, stimulation of either receptor resulted in a cross-desensitization of the subsequent response to the other agonist. Treatment of oocytes with either H-8 (100 μM), staurosporine (400 nM), okadaic acid (200 nM), phorbol myristate acetate (5 nM) or forskolin (50 μM) plus phosphodiesterase inhibitor did not affect either the desensitization or the agonist-evoked inhibition of Ca²⁺ channels.
5. These results suggest that the rate of rapid desensitization is dependent on the α₁ subtype of the neuronal Ca²⁺ channel, and that a common phosphorylation-independent mechanism underlies the heterologous desensitization between opioid receptor subtypes.

     

High prevalence of hepatitis C virus infection in schistosomiasis japonica patients associated with hepatocellular carcinoma

Schistosomiasis japonica (SCJ) patients frequently develop hepatocellular carcinoma (HCC). This study investigated relationship between SCJ infection, hepatitis virus infection, and incidence of HCC, in 25 patients with chronic SCJ infection and HCC (SCJ with HCC group), 51 patients with chronic SCJ infection without HCC (SCJ group) and 65 HCC patients without SCJ (HCC group). Number of patients who were positive to HBsAg or hepatitis B virus DNA were 4 (16.0%) in the SCJ with HCC group, none (0%) in the SCJ group, and 5 (7.9%) in the HCC group; while number of patients who were positive to anti-hepatitis C virus antibody were 21 (87.5%) in the SCJ with HCC group, 3 (5.9%) in the SCJ group, and 58 (84.6%) in the HCC group. Biopsy was performed for all patients, and background histological features of each specimen were evaluated based on the histological activity index scoring system. Mean scores of inflammatory changes in both the portal area and parenchyma in the SCJ with HCC group were significantly higher than those in the SCJ group. Nodular formation which is common in post-viral hepatitis was frequently observed in the SCJ with HCC group, and histological changes in non-cancerous area of the SCJ with HCC group showed the characteristics of chronic viral hepatitis. We conclude that infection of hepatitis virus, particularly hepatitis C virus, affects synergistically on the hepatocarcinogenesis in patients having SCJ infection.     

Accumulation of L-[2-(F-18)]fluorophenylalanine in peri-infarct area in a patient with acute cerebral infarction

We studied the brain uptake of amino acid in a patient with acute cerebral infarction with L-[2-(F-18)] fluorophenylalanine and positron emission tomography. The increased accumulation of the ligand was specifically found in the peri-infarct area where oxygen metabolism was still maintained but decreased later in the 72-day follow-up period. The kinetic analysis revealed that increased accumulation was not due to increased transport from the blood to the brain but to delayed washout from the brain to the blood. Although the mechanism is still unknown, abnormally high accumulation of L-[F-18]fluorophenylalanine may predict delayed neuronal changes after ischemic insults of the brain.     

Multiple tumor-suppressor-1 gene and esophageal-carcinoma

The multiple tumor suppressor 1 (MTS1) gene is homozygously deleted frequently in cell lines derived from a wide variety of tumors. We investigated the deletion of the MTS1 gene in esophageal cancer cell lines and primary esophageal squamous carcinomas using the polymerase chain reaction. Sixteen and 15 of 23 esophageal cancer cell lines showed homozygous deletion of MTS1 exon 1 and exon 2, respectively, while none of 21 primary esophageal carcinomas showed the deletion. An analysis of MTS1 gene mutations was carried out by direct DNA sequencing in 8 cell lines and 21 primary carcinomas showing no homozygous deletion. In contrast to previous reports of esophageal carcinoma, there were no mutations recognized in the region sequenced. Our study suggests that the inactivation of the MTS 1 gene may play an important role in esophageal carcinoma cell lines but may be less important in primary carcinomas of the human esophagus.     

In utero methylmercury exposure differentially affects the activities of selenoenzymes in the fetal mouse brain

Pregnant ICR mice were subcutaneously injected with 0,5, or 3x3 mg Hg/kg of methylmercury (MeHg) on days 12,13, and 14(G12-14) of gestation and were sacrificed on G17. Activity of selenoenzymes, including glutathione peroxidase (GPx) and 5'- or 5-iodothyronine deiodinases (5'-DI, 5-DI), was determined in fetal brain and placenta. MeHg did not affect the concentration of Se in these tissues, while it significantly inhibited the activity of GPx in the fetal brain and placenta, but not in the maternal brain. Although the levels of thyroid hormones in the maternal and fetal plasma were not affected by MeHg, 5-DI decreased and 5'-DI increased in the fetal brain, as if they had responded to hypothyroidism. Because the level of T4 in the fetal plasma was not affected by MeHg, these changes in enzymatic activities may result in a harmful excess of T3 in the fetal brain. In addition, 5-DI activity was increased in the placenta of MeHg-treated mice. These effects of prenatal MeHg exposure on fetal and placental DIs differed from those of dietary-induced Se deficiency, where the activities of DIs were decreased or not affected. Further evaluation of the effect of MeHg on selenoenzymes, especially 5-DIs, is warranted.     

Poorer nutritional status of the older-generation in a Japanese farming village, as related to fewer types of food consumed

This study was carried out to determine whether the most recent nutritional improvements in Japanese farming villages were due to improvement in the diet of the young only or across all ages. Food duplicates for 24 h were collected. The number of food items and the adequacy of each nutrient level were compared between subgroups, by age of cooking and those eating meals. The older-generation used and ate fewer food items resulting in poorer nutrition. The most recent improvement in nutrition in the farming villages of Japan was found mainly to affect the younger-generation with the older-generations being more poorly nourished.