Role of ionotropic glutamatergic and GABAergic inputs on the firing activity of neurons in the external pallidum in awake monkeys

The neurons in the external segment of the pallidum (GPe) in awake animals maintain a high level of firing activity. The level and pattern of the activity change with the development of basal ganglia disorders including parkinsonism and hemiballism. The GPe projects to most of the nuclei in the basal ganglia. Thus exploring the mechanisms controlling the firing activity is essential for understanding basal ganglia function in normal and pathological conditions. To explore the role of ionotropic glutamatergic and GABAergic inputs to the GPe, unit recordings combined with local injections of receptor antagonists were performed in awake monkeys. Observations on the effects of local application of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/kainate antagonist 1,2,3,4-tetrahydro-6-nitro-2, 3-dioxo-benzo[f]quinoxaline-7-sulfonamide, the N-methyl-D-aspartic acid (NMDA) antagonist 3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid, and the GABAA antagonist gabazine as well as the effects of muscimol blockade of the subthalamic nucleus on the spontaneous firing rate, firing patterns, and cortical stimulation induced responses in the GPe suggested the following: sustained glutamatergic and GABAergic inputs control the level of the spontaneous firing of GPe neurons; both AMPA/kainate and NMDA receptors are activated by glutamatergic inputs; some GPe neurons receive glutamatergic inputs originating from areas other than the subthalamic nucleus; no GPe neurons became silent after a combined application of glutamate and GABA antagonists, suggesting that GPe neurons have intrinsic properties or nonionotropic glutamatergic tonic inputs that sustain a fast oscillatory firing or a combination of a fast and a slow oscillatory firing in GPe neurons.     

Constitutive Expression of a Bacterial Pattern Recognition Receptor, CD14, in Human Salivary Glands and Secretion as a Soluble Form in Saliva

Saliva contains a number of proteins and glycoproteins that protect oral tissues, but little is known about the role of human saliva in innate immunity. Here we showed that human major salivary gland cells constitutively expressed a bacterial pattern recognition receptor, CD14, by immunohistochemistry. Human salivary gland cells in culture express CD14 mRNA and a 55-kDa CD14 protein in, but not on the cells, and secrete a soluble form with the same molecular mass. Human whole saliva contains a 55-kDa CD14, and the concentration of parotid saliva was 10-fold higher than whole saliva, which is comparable to that of serum CD14. Levels of CD14 in unstimulated whole and parotid saliva were unchanged before and after a meal and between unstimulated and stimulated saliva, indicating that saliva CD14 is constitutively secreted into the oral cavity. In contrast, lipopolysaccharide (LPS)-binding protein was below the detectable level. The saliva CD14 is functionally active in that it mediated the activation of CD14-lacking intestinal epithelial cells by LPS in a Toll-like receptor 4-dependent manner. These results suggested that saliva CD14 is important for the maintenance of oral health and possibly intestinal homeostasis.     

Frequent co-localization of Cox-2 and laminin-5 gamma2 chain at the invasive front of early-stage lung adenocarcinomas

Laminin-5 is an extracellular matrix protein that plays a key role in cell migration and tumor invasion. Cox-2 is an induced isoform of cyclooxygenases that plays an important role in carcinogenesis, suppression of apoptosis, angiogenesis, and metastasis of colon cancer. We report frequent co-expression of cox-2 and laminin-5 at the invasive front of early-stage lung adenocarcinomas. We investigated the expression of cox-2 and laminin-5 immunohistochemically in 102 cases of small-sized lung adenocarcinoma (maximum dimension, 2 cm or less). Cox-2 and laminin-5 were expressed in 97 (95.1%) and 82 (80.4%) cases, respectively. Both were preferentially localized in cancer cells at the cancer-stroma interface, although cox-2 tended to show a diffuse staining pattern in some cases. A comparison of their staining patterns revealed a striking similarity in their distribution in 24 cases, and a partial overlap between their localization in another 20 cases. Moreover, an overall correlation was found between the expression levels of cox-2 and laminin-5 (P = 0.018). To gain insight into the mechanisms that regulate the expression of these proteins, we additionally studied their expression in 58 cases of stage I lung adenocarcinoma, in which p53 status was determined by immunohistochemistry, polymerase chain reaction-single strand conformation polymorphism analysis, and direct sequencing. The results showed that tumors with mutant p53 tended to express more cox-2 than those with wild-type p53 (P = 0.080). Also, tumors that overexpressed p53 had higher levels of cox-2 and laminin-5 than those without p53 overexpression (P = 0.032 and 0.047, respectively). Further immunohistochemical analysis showed that tumors that overexpressed both epidermal growth factor receptor (EGFR) and erbB-2 had higher levels of cox-2 and laminin-5 than those without concomitant overexpression of these proteins (P = 0.014 and P = 0.018, respectively). To see whether EGFR signaling is involved in cox-2 and laminin-5 expression, we further conducted in vitro analyses using six lung adenocarcinoma cell lines (A549, HLC-1, ABC-1, LC-2/ad, VMRC-LCD, and L27). Western blot analyses showed that cox-2 mRNA levels, and to a lesser extent laminin-5 gamma2 mRNA levels, correlated with the expression levels of erbB-2 and the phosphorylated form of MAPK/ERK-1/2 protein. The addition of transforming growth factor-alpha increased both cox-2 and laminin-5 gamma2 mRNA levels in A549, ABC-1, and L27 with different kinetics; the induction of cox-2 occurred earlier than that of laminin-5 gamma2. Finally, the migration of ABC-1 cells was inhibited by MAP kinase kinase inhibitor PD98059 and a selective cox-2 inhibitor NS-398. In contrast, the migration of A549 cells was inhibited by PD98059, but much less effectively by NS-398. These results suggest that co-stimulatory mechanisms may exist that increase the expression of cox-2 and laminin-5 at the invasive front of lung adenocarcinomas and that EGFR signaling could be one of the mechanisms. Further investigations are warranted concerning the role of cox-2 and laminin-5 in cancer cell invasion and the significance of p53 and EGFR signaling in the regulation of cox-2 and laminin-5 expression.     

Clinical heterogeneity of neonatal intrahepatic cholestasis caused by citrin deficiency: case reports from 16 patients

A deficiency of citrin, which is encoded by the SLC25A13 gene, causes both adult-onset type II citrullinemia (CTLN2) and neonatal intrahepatic cholestasis (NICCD). We analyzed 16 patients with NICCD to clarify the clinical features of the disease. Severe intrahepatic cholestasis with fatty liver was the most common symptom, but the accompanying clinical features were variable, namely; suspected cases of neonatal hepatitis or biliary atresia, positive results from newborn screening, tyrosinemia, failure to thrive, hemolytic anemia, bleeding tendencies and ketotic hypoglycemia. Laboratory data showed elevated serum bile acid levels, hypoproteinemia, low levels of vitamin K-dependent coagulation factors, and hypergalactosemia. Hypercitrullinemia was detected in 11 out of 15 patients examined. Most of the patients were given a lactose-free and/or medium chain triglycerides-enriched formula and lipid-soluble vitamins. The prognosis of the 16 patients is going fairy well at present, but we should observe these patients carefully to see if they manifest any symptom of CTLN2 in the future.     

Massive apoptosis in infantile myofibromatosis. A putative mechanism of tumor regression.

Two cases of solitary infantile myofibromatosis (IM) are presented. Solitary IM are tumors prone to spontaneous regression. Histopathologically, several tumor lobules in our IM cases had central areas of massive cell death, with nuclear pyknosis, cytoplasmic hyalinization and nuclear fragmentation but without lymphoid or neutrophilic cell infiltration. These central cell death areas consisted of about 40% in case 2 and 50% in case 1 of the entire tumor tissues, respectively. Electron microscopy revealed that the condensed nuclei and cytoplasm were fragmented into "apoptotic bodies", with or without phagocytosis by histiocytes. DNA fragmentation, as evidenced by the terminal deoxy transferase-mediated uptake of biotinylated dUTP, was identified at massive cell death areas on paraffin sections from both cases. A characteristic 180- to 190-bp nucleosomal ladder was detected in DNA obtained from the tumor cells in case 1. The collective evidence suggested that these tumors underwent a central, massive apoptosis. As massive cell death similar to that seen in the present cases has been described in other documented cases of IM, we propose that the spontaneous regression that frequently occurs with this type of tumor may be mediated by massive apoptotic cell death.     

An Epstein-Barr virus-producer line Akata: Establishment of the cell line and analysis of viral DNA

An Epstein-Barr virus (EBV)-producer line, designated Akata, was established from a Japanese patient with Burkitt's lymphoma. The Akata line possessed the Burkitt's-type chromosome translocation, t(8q-; 14q+), and was derived from the tumor cell. Akata cells produced a large quantity of transforming virus upon treatment of cells with anti-immunoglobulin antibodies (Takada, 1984). Southern blot analysis of viral DNA indicated that the Akata EBV is nondefective and more representative of wild-type viruses. Akata cells should be useful as a source of EBV.     

Topographical projections from the thalamus to the putamen in the cat

Thalamic projections to the putamen (Put) in the cat were studied by the retrograde horseradish peroxidase method. Major thalamic projections to the Put originate from the midline and intralaminar nuclear regions including the centre médian-parafascicular complex (CM-Pf). The other thalamic projections to the Put arise mainly from the suprageniculate nucleus (Sg), magnocellular division of the medial geniculate nucleus (MGm), caudomedial part of the lateroposterior nucleus (LP) and ventrolateral part of the ventromedial nucleus (VM). The VM projects to the rostral Put, while the posterior thalamic regions (Sg, MGm, LP) project to the caudal Put.     

Subchronic exposure to arsenic through drinking water alters expression of cancer-related genes in rat liver

Although arsenic exposure causes liver disease and/or hepatoma, little is known about molecular mechanisms of arsenic-induced liver toxicity or carcinogenesis. We investigated the effects of arsenic on expression of cancer-related genes in a rat liver following subchronic exposure to sodium arsenate (1, 10, 100 ppm in drinking water), by using real-time quantitative RT-PCR and immunohistochemical analyses. Arsenic accumulated in the rat liver dose-dependently and caused hepatic histopathological changes, such as disruption of hepatic cords, sinusoidal dilation, and fatty infiltration. A 1-month exposure to arsenic significantly increased hepatic mRNA levels of cyclin D1 (10 ppm), ILK (1 ppm), and p27(Kip1) (10 ppm), whereas it reduced mRNA levels of PTEN (1 ppm) and beta-catenin (100 ppm). In contrast, a 4-month arsenic exposure showed increased mRNA expression of cyclin D1 (100 ppm), ILK (1 ppm), and p27(Kip1) (1 and 10 ppm), and decreased expression of both PTEN and beta-catenin at all 3 doses. An immunohistochemical study revealed that each protein expression accords closely with each gene expression of mRNA level. In conclusion, subchronic exposure to inorganic arsenate caused pathological changes and altered expression of cyclin D1, p27(Kip1), ILK, PTEN, and beta-catenin in the liver. This implies that arsenic liver toxicity involves disturbances of some cancer-related molecules.