Astroglial responses against Abeta initially occur in cerebral primary cortical cultures: species differences between rat and cynomolgus monkey.

In the present study, we investigated how amyloid beta (Abeta) peptides initially affect neuronal cells in primary cerebral cortical cultures from rat and cynomolgus monkey. In these cultures, complicated interactions between glial and neuronal cells occur; moreover, synaptic interactions similar to those observed in vivo also occur between neuronal cells in these cultures. In this study, we applied low concentrations of Abeta to these well-characterized primary cultures to investigate how Abeta initially affects neurons or astroglial cells. In both rat and monkey cortical cultures, treatment with low concentrations of Abeta failed to drastically change or damage of neurons. Abeta treatment, however, significantly activated astrocytes, resulting in increased apolipoprotein E (ApoE) production. Rat astrocytes were more sensitive to Abeta than monkey astrocytes, and responded to Abeta via a different mechanism. In monkey astrocyte cultures, only direct treatment with Abeta increased ApoE production. In rat astrocyte cultures, however, treatment with conditioned media from cortical cultures grown with Abeta increased ApoE production, indicating that some sort of neuron-derived soluble factor(s) was also involved in activating rat astrocytes. These species differences suggest that monkey cortical cultures would be more useful as an in vitro model system to understand the details of how Abeta accumulates in the human brain, since monkeys are phylogenetically more similar to humans.     

Isolation and characterization of a highly divergent HIV-2[GH-2]: generation of an infectious molecular clone and functional analysis of its rev-responsive element in response to primate retrovirus transactivators (Rev and Rex).

A highly divergent HIV-2 designated as HIV-2[GH-2] was obtained from an AIDS-related complex (ARC) patient in Ghana. A full-length molecular clone of this isolate was obtained and a biologically active clone was constructed. Its restriction pattern differed from that of prototype HIV-2[GH-1] in 25 of 35 restriction sites, but was strikingly similar to a previously characterized HIV-2 isolate from a Ghanaian (HIV-2ALT). The conserved integrase region (288-bp fragment) previously displayed 95% identity with that of ALT but 17-20% divergence from the HIV-2 prototype member, and a new distinct subgroup (HIV-2b) of HIV-2 consisting of GH-2 and ALT was postulated (Miura et al. 1991.) These isolates, however, were biologically distinguishable from each other by its replication capacity in a monocyte line, U937, in which GH-2 could not grow but ALT grew well. In addition, the nucleotide sequence of the LTR of this new isolate displays 21% divergence from that of prototype HIV-2[GH-1], but the core enhancer, Sp1 binding sites and TATA box were conserved. Although the 3' half of the env gene sequence which is deleted in HIV-2ALT clone showed 27% diversity from the prototype, functional differences in the rev-responsive element were not observed.     

Sequential change of the hypervariable region of the hepatitis C virus genome in acute infection

Hepatitis C virus (HCV) infection is characterized by persistence of liver inflammation that often leads to end-stage liver disease, although the mechanisms are not fully understood. A hyper-variable region (HVR) has been reported in the E2/NS1 region of the HCV genome, in which striking diversity is found among different HCV isolates. To investigate the association of the HVR alterations with the clinical courses of HCV infection, a longitudinal analysis of the HVR in patients with acute HCV infection was carried out. Plasma samples were obtained at several times in three patients with acute hepatitis C. Plasma RNA was extracted and reverse transcribed, and DNA fragments that included the HVR were amplified by PCR. The sequences of the HVR were directly determined from the PCR products by the dideoxy chain termination method, from which amino acid sequences were deduced. In all cases, plasma HCV-RNA disappeared with the improvement of the initial alanine aminotransferase (ALT) elevation, but HCV-RNA reappeared about 1 year later with or without deterioration of the hepatitis. In a case of sporadic acute hepatitis, the HCV in the recurrent phase had seven amino acid substitutions in the HVR compared with that in the acute phase, although no amino acid changes were noted during the initial acute phase. In a case of posttransfusion hepatitis, a marked difference was observed between the acute and the recurrent phases, with an amino acid homology of 30% (8/27). The mutation rate of the HVR had a tendency to accelerate as the HCV infection progressed to the chronic stage. In conclusion, the HVR changes serially during the course of acute HCV infection, and these HVR changes may play a part in the chronicity of HCV infection. © 1994 Wiiey-Liss, Inc.     

Anterior urethral recurrence of superficial bladder cancer: its clinical significance

The aim of this study was to reveal the clinical features of anterior urethral recurrence in patients with superficial bladder cancer, and to determine the appropriate treatment. Three hundred and three patients with superficial bladder cancer, who were newly diagnosed and initially treated conservatively in our hospital between 1965 and 1990, were followed for at least 5 years and their clinical outcomes were analyzed. Clinical factors, including anterior urethral recurrence, were evaluated statistically regarding tumor progression. Eight patients (2.6%) had anterior urethral recurrence following superficial bladder cancer. Twenty-four patients (7.9%) had tumor progression and 149 (49.2%) had tumor recurrence. In a multivariate analysis using a logistic model, anterior urethral recurrence was the most important factor, followed by histological grade. Four of 5 patients who were treated for anterior urethral recurrent tumors by transurethral resection showed progression and died of the cancer within one year. Two of the remaining three patients who underwent radical cysto-urethrectomy at the time of anterior urethral recurrence survived. Anterior urethral recurrence following superficial bladder cancer is a predictor for rapid subsequent malignant progression. Once there is anterior urethral recurrence, radical intensive therapy, including radical cysto-urethrectomy, should be carried out immediately.     

Loss of Hrs in the Central Nervous System Causes Accumulation of Ubiquitinated Proteins and Neurodegeneration

The endosomal sorting complex required for transport (ESCRT) proteins form multimolecular complexes that control multivesicular body formation, endosomal sorting, and transport ubiquitinated membrane proteins (including cell-surface receptors) to the endosomes for degradation. There is accumulating evidence that endosomal dysfunction is linked to neural cell degeneration in vitro, but little is known about the relationship between neural disorders and ESCRT proteins in vivo. Here we specifically deleted the hrs gene, ESCRT-0, in the neurons of mice by crossing loxP-flanked hrs mice with transgenic mice expressing the synapsin-I Cre protein (SynI-cre). Histological analyses revealed that both apoptosis and a loss of hippocampal CA3 pyramidal neurons occurred in the hrs^flox/flox;SynI-cre mice. Notably, the hrs^flox/flox;SynI-cre mice accumulated ubiquitinated proteins, such as glutamate receptors and an autophagy-regulating protein, p62. These molecules are particularly prominent in the hippocampal CA3 neurons and cerebral cortex with advancing age. Accordingly, we found that both locomotor activity and learning ability were severely reduced in the hrs^flox/flox;SynI-cre mice. These data suggest that Hrs plays an important role in neural cell survival in vivo and provide an animal model for neurodegenerative diseases that are known to be commonly affected by the generation of proteinaceous aggregates.     

Modifying effects of 1'-acetoxychavicol acetate (ACA) and the novel synthetic retinoids Re-80, Am-580 and Am-55P in a two-stage carcinogenesis model in female rats

Effects of dietary administration of 1'-acetoxychavicol acetate (ACA) and the novel synthetic retinoids 4-[1-hydroxy-3-oxo-3-(5,6,7,8-tetrahydro-3-hydroxy-5,5,8,8-tetramethyl-2-naphthalenyl)-1-propenyl]benzoic acid (Re-80); 4-[(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)carboxamido]benzoic acid (Am-580); and 6-[(3,5-di-tert-butylphenyl) carbamoyl]nicotinic acid (Am-55P) were examined using a two-stage rat carcinogenesis model. A total of 190 female SD rats was treated sequentially with 1,2-dimethylhydrazine (DMH, s.c.); 7,12-dimethylbenz(a)anthracene (DMBA, i.g.); and 2,2'-dihydroxy-di-n-propylnitrosamine (DHPN, in the drinking water) during the first three weeks (DDD-initiation), and an additional 60 rats received the vehicle alone (non-initiation). One week after the completion of the initiation period, they were divided into nine groups and administrated Re-80 (at dose levels of 1.0 or 0.4 ppm), Am-580 (20 or 4 ppm), Am-55P (20 ppm), ACA (100 ppm), all-trans-retinoic acid (10 or 2 ppm) or no supplement in the diet for 33 weeks, until survivors were euthanatized at week 37 weeks. After DDD-initiation, all-trans-retinoic acid at the high dose delayed the development of mammary tumors. The multiplicity of colon tumors in the group fed Am-55P and the incidences of nephroblastomas with ACA or Am-580 were decreased as compared with the control values, but the other chemicals had no modifying effects on tumor development in any organs. Thus, among ACA and the novel synthetic retinoids tested, only Am-55P showed a weak inhibitory effect on a neoplasm of general interest under the present experimental conditions.     

Suppression of UV-induced mutations by wild-type p53 protein in human osteosarcoma cells

We have examined whether the tumour suppressor p53 protein suppressedUV-induced mutations in the hypoxathine-guanine phosphoribosyltransferase (HPRT) gene and in the supF gene of the shuttlevector plasmid pMY189. We used human osteosarcoma Saos-LP12cells, in which wild type (wt) p53 protein was induced by treatmentwith isopopyl-(ß-D-thiogalactopyranoside. The inductionof wt p53 protein suppressed UV-induced mutations but not spontaneousmutations in the HPRT gene. The frequency of UV-induced mutationsinduced by UV-irradiation of the plasmid was also significantlylower in cells with induced wt p53 protein than in the uninducedcells. In addition, we found that frequency of G : C to A :T transition mutations which occurred at the 3' base pair ofdipyrimidine sites were significantly lower in the cells withinduced wt p53 protein than in the uninduced cells. These findingssuggestthat wt p53 protein may play roles in modulating DNArepair pathway, resulting in the suppression of UV-induced mutations. ¹To whom correspondence should be addressed     

p38 Mitogen-activated protein kinase activation in endothelial cell is implicated in cell alignment and elongation induced by fluid shear stress.

Fluid shear stress is thought to be important in maintaining the phenotype of endothelial cells (ECs) in vivo. The purpose of the study was to determine the effect of varying levels of laminar shear stress on EC elongation and alignment and the role of p38 mitogen-activated protein kinase (MAPK) on the morphologic change induced by shear stress. Cultured bovine aortic ECs were subjected to 1, 4, 7, 14, or 20 dyne/cm(2) laminar steady shear stress. On morphometric analysis of static ECs, the average orientation angle was 41 degrees , whereas after 24 h shear stress at 1, 4, 7, 14, and 20 dyne/cm(2) the angles were 34 degrees, 33 degrees, 16 degrees, 11 degrees, and 10 degrees, respectively. The shape index of static ECs was 0.76, whereas the indexes of ECs exposed to shear stress were 0.72, 0.72, 0.65, 0.50, and 0.47, respectively. The time and the magnitude of activation of p38 MAPK were dependent on the level of shear stress. The results indicate that a minimum shear stress of 7 to 14 dynes/cm(2) is necessary for cell alignment and elongation and this correlates with activity of p38 MAPK. ECs exposed to shear stress in the presence of the p38 MAPK inhibitor SB-203580 did not orient in any manner and the shape index was similar to the static cells.     

PRIMARY LEIOMYOSARCOMA OF BONE

A 62-year-old female with primary leiomyosarcoma of the left femur is reported with a review of 21 cases reported in the literature. The resected specimen showed that the tumor extended from the femoral head to the diaphysis for 13cm in length. The tumor showed mainly intramedullary proliferation, but extraosseous growth was also noted at the great trochanter. Microscopic examination revealed well differentiated leiomyosarcoma characterized by interlacing bundles of fusiform cells with eosinophilic cytoplasm and rod-shaped hyperchromatic nuclei. PAP stain of actin on the tumor cells was positive. On electron microscopy, microfilament of 6–8 nm in diameter, dense bodies, plnocytotic vesicles, marginal attachment plate, and basal lamina were noted. The patient died with pulmonary metastasis, 1 year and 7 months after the operation. An autopsy showed metastases in the right pelvic cavity and bilateral lungs, and confirmed the primary site to be the left femur.