Immunological detection of severe acute respiratory syndrome coronavirus by monoclonal antibodies

In order to establish immunological detection methods for severe acute respiratory syndrome coronavirus (SARS-CoV), we established monoclonal antibodies directed against structural components of the virus. B cell hybridomas were generated from mice that were hyper-immunized with inactivated SARS-CoV virion. By screening 2,880 generated hybridomas, we established three hybridoma clones that secreted antibodies specific for nucleocapsid protein (N) and 27 clones that secreted antibodies specific for spike protein (S). Among these, four S-protein specific antibodies had in vitro neutralization activity against SARS-CoV infection. These monoclonal antibodies enabled the immunological detection of SARS-CoV by immunofluorescence staining, Western blot or immunohistology. Furthermore, a combination of monoclonal antibodies with different specificities allowed the establishment of a highly sensitive antigen-capture sandwich ELISA system. These monoclonal antibodies would be a useful tool for rapid and specific diagnosis of SARS and also for possible antibody-based treatment of the disease.     

Troglitazone induces p27Kip1-associated cell-cycle arrest through down-regulating Skp2 in human hepatoma cells

Increasing evidence has confirmed that ligands for peroxisome proliferator-activated receptor gamma (PPARgamma) exhibit antitumoral effects through inhibition of cell proliferation and induction of cell differentiation in several malignant neoplasms. Recently, we have documented the accumulation of a cyclin-dependent kinase inhibitor, p27(Kip1), as well as an unexpected accumulation in cyclin E in G1-arrested human hepatoma cells treated with the PPARgamma ligand troglitazone. Simultaneous accumulations in both p27(Kip1) and cyclin E are known to be characteristic phenotypes in cells derived from mice lacking Skp2, an F-box protein component of the SCF ubiquitin-ligase complex. Thus, the aim of the present study was to assess whether Skp2 might be involved in the down-regulation of p27(Kip1) in troglitazone-treated human hepatoma cells. A striking decrease in Skp2 expression and a reciprocal increase in p27(Kip1) expression were found in troglitazone-treated hepatoma cells but not in those cells treated with other PPARgamma ligands such as pioglitazone and ciglitazone. Quantitative real-time RT-PCR analysis showed that troglitazone down-regulated Skp2 at the mRNA levels. Consistently, ectopic overexpression in Skp2 brought resistance to troglitazone, resulting in a decreased population of arrested cells at the G1 phase compared with that in the mock-transfected cells. In surgically resected hepatocellular carcinoma (HCC) tissue, an increased expression in Skp2 was found in both the moderately differentiated HCCs and the poorly differentiated HCCs. In conclusion, troglitazone attenuated Skp2 expression, thereby promoting p27(Kip1) accumulation in human hepatoma cells. This therapeutic potential of the ligand may lead to new cell-cycle-based antitumor strategies for advanced HCCs.     

Suppression of adjuvant-induced arthritic bone destruction by cyclooxygenase-2 selective agents with and without inhibitory potency against carbonic anhydrase II.

In vitro assays revealed that COX-2 inhibitors with CA II inhibitory potency suppressed both differentiation and activity of osteoclasts, whereas that without the potency reduced only osteoclast differentiation. However, all COX-2 inhibitors similarly suppressed bone destruction in adjuvant-induced arthritic rats, indicating that suppression of osteoclast differentiation is more effective than that of osteoclast activity for the treatment. INTRODUCTION: Cyclooxygenase (COX)-2 and carbonic anhydrase II (CA II) are known to play important roles in the differentiation of osteoclasts and the activity of mature osteoclasts, respectively. Because several COX-2 selective agents were recently found to possess an inhibitory potency against CA II, this study compared the bone sparing effects of COX-2 selective agents with and without the CA II inhibitory potency. MATERIALS AND METHODS: Osteoclast differentiation was determined by the mouse co-culture system of osteoblasts and bone marrow cells, and mature osteoclast activity was measured by the pit area on a dentine slice resorbed by osteoclasts generated and isolated from bone marrow cells. In vivo effects on arthritic bone destruction were determined by radiological and histological analyses of hind-paws of adjuvant-induced arthritic (AIA) rats. RESULTS: CA II was expressed predominantly in mature osteoclasts, but not in the precursors. CA II activity was inhibited by sulfonamide-type COX-2 selective agents celecoxib and JTE-522 similarly to a CA II inhibitor acetazolamide, but not by a methylsulfone-type COX-2 inhibitor rofecoxib. In vitro assays clearly revealed that celecoxib and JTE-522 suppressed both differentiation and activity of osteoclasts, whereas rofecoxib and acetazolamide suppressed only osteoclast differentiation and activation, respectively. However, bone destruction in AIA rats was potently and similarly suppressed by all COX-2 selective agents whether with or without CA II inhibitory potency, although only moderately by acetazolamide. CONCLUSIONS: Suppression of osteoclast differentiation by COX-2 inhibition is more effective than suppression of mature osteoclast activity by CA II inhibition for the treatment of arthritic bone destruction.     

Fetal germinal matrix and intraventricular hemorrhage associated with periventricular leukomalacia]

Subependymal germinal matrix hemorrhage with intraventricular hemorrhage(SEIVH) is a common complication associated with delivery in preterm neonates but has rarely been observed in the fetus. We report a fetus with SEIVH, hydrocephalus and periventricular leukomalacia(PVL). Although this fetus had uneventful prenatal periods, transabdominal ultrasound examination(US) at 33 weeks of pregnancy revealed SEIVH and hydrocephalus, and MRI at 36 weeks did associated PVL. While no events reported that could explain the onset of SEIVH, PVL was considered to be the results of anoxic events associated with SEIVH. In addition to US, information provided by fetal MRI, especially T 2-weighted image, permits a better understanding of the pathophysiology of fetal SEIVH with PVL.     

Lysosomal enzyme activities in cultured lymphoid cell lines

Several lysosomal enzyme activities in cultured lymphoid cell lines were studied during 3 phases of cell culture; logarithmic growth phase, stationary phase and decline phase.Enzyme induction during cell growth was found in N-acetyl-hexosaminidase, β-galactosidase and α-l-fucosidase, but no induction in a-d-mannosidase, α-glucosidase and β-glucuronidase. The latter two enzymes were unchanged during all cell culture phases.A drop in α-l-fucosidase and α-d-mannosidase activity was found during the stationary and decline phases of cell culture.     

Lymphangioma of the parotid gland

Parotid gland tumors in infancy and childhood are rarely seen and infrequently documented. Among them, hemangioendothelioma and mixed tumors are the most common. Parotid lymphangioma is very rare. This report presents a case of parotid lymphangioma and discusses the clinical, gross, and histological features, differential diagnosis, and treatment.     

Convergence of afferent inputs from the chorda tympani, lingual-tonsillar and pharyngeal branches of the glossopharyngeal nerve, and superior laryngeal nerve on the neurons in the insular cortex in rats

The responses of single neurons in the insular cortex to electrical stimulation of the chorda tympani (CT), lingual-tonsillar branch of the glossopharyngeal (LT-IXth) nerve, pharyngeal branch of the glossopharyngeal (PH-IXth) nerve, and superior laryngeal (SL) nerve were recorded in anaesthetized and paralyzed rats. Ninety-four neurons responding to stimulation of at least one of the four nerves were identified from the insular cortex. Most of the neurons were located in the posterior portion of the insular cortex; the mean location was 0.8 mm anterior to the anterior edge of the joining of the anterior commissure (AC) and was 1.4 mm dorsal to the rhinal fissure (RF). Of the 94 neurons, 84 (89%) received convergent inputs from two or more nerves, and the remaining 10 (11%) received inputs from one nerve. The neurons responding to the CT stimulation were distributed more anteriorly than those responding to other three nerves in the anterior-posterior dimension. Our results indicate that the neurons recorded mainly from the posterior portion of the insular cortex receive convergent inputs from the oropharyngolaryngeal regions.     

Neurochemical correlates of selective neuronal loss following cerebral ischemia: role of decreased Na, K-ATPase activity

In order to investigate the role of Na+,K(+)-ATPase in the development of neuronal necrosis following cerebral ischemia, ischemia was induced in gerbils by occluding the common carotid artery unilaterally for 10 min. A time-course analysis revealed that significant reductions of the Na+,K(+)-ATPase activity in the cerebral cortex and hippocampus were manifested at 15 min, 30 min, and 1 h, and returned to the control level one day following recirculation. No apparent alterations of the Mg(2+)-ATPase activity, on the other hand, were obtained throughout the experimental period. Furthermore, Scatchard analyses of [3H]ouabain binding to the cerebral cortex membranes disclosed that the Bmax values invariably decreased without any change of Kd values following ischemia. It has also been shown that treatment of the animals with an agent known to mitigate ischemic neuronal necrosis, i.e. BY-1949, significantly reversed such derangements. These results suggest that the recovery of decreased Na+,K(+)-ATPase activity shortly after ischemia exerts a protective effect against ischemic brain damage.