Comparison of enzyme phenotypes in human bladder tumours and experimentally induced hyperplastic and neoplastic lesions of the rat urinary bladder. A combined histochemical and immunohistochemical approach

The expression of a number of enzymes involved in drug metabolism, membrane function etc. was compared in hyperplastic and neoplastic lesions of the rat bladder and in human bladder tumours. Transitional cell carcinomas (TCC) in both rat and Man were characterized by decreased alkaline phosphatase (ALP) and increased gamma-glutamyl transpeptidase (GGT), beta-glucuronidase (beta-G1), succinate dehydrogenase (SD) and glucose-6-phosphate dehydrogenase (G6PD) activities. In addition, binding for antibodies specific for different cytochrome P-450 species (UT50, PB3a, MC1, MC2) and microsomal epoxide hydrolase (mEHb) was elevated in both murine and human tumours. Comparison of the enzyme phenotype in hyperplastic lesions induced by freeze ulceration or uracil administration with that in preneoplastic papillary or nodular hyperplasia (PNH) and TCC suggested, however, that most of the alteration in enzyme content or activity was non-specific and related to requirements for epithelial cell proliferation. On the other hand, the decreased ALP, and increased GGT and beta-G1 activity appeared more directly related to neoplastic transformation. The results suggested that qualitative differences exist between reactive hyperplasia and preneoplastic or neoplastic lesions in the urinary bladder. The finding of increased cytochrome P-450, in clear contrast to the reduction characteristic of preneoplastic hepatic lesions, may be important with regard to the observed difference in neoplastic transformation between the bladder and liver in response to drug metabolising enzyme inducers.     

Correlative histochemical studies on preneoplastic and neoplastic lesions in the kidney of rats treated with nitrosamines

Renal tubular lesions induced in male rats by two different carcinogens, N-nitrosomorpholine (NNM) and N-ethyl-N-hydroxyethylnitrosamine (EHEN), using a limited exposure "stop" protocol were investigated histochemically to demonstrate phenotypic cellular changes. The parameters measured included basophilia, glycogen content and the activity of the enzymes glucose-6-phosphatase (G6PASE), glycogen synthetase (SYN), glycogen phosphorylase (PHO), glucose-6-phosphate dehydrogenase (G6PDH), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), succinate dehydrogenase (SDH), alkaline phosphatase (ALP), acid phosphatase (ACP) and gamma-glutamyl transpeptidase (gamma-GT). The lesions observed were predominantly of either basophilic or oncocytic types. In each case, tubular lesions (altered tubules) appeared to give rise to epithelial tumors (epitheliomas) with the same cellular phenotype. Basophilic tubules and epitheliomas proved to be strongly positive for GAPDH and G6PDH while demonstrating a reduction or loss of G6PASE, ALP, ACP, gamma-GT, and SDH compared with controls and the surrounding proximal or distal tubules. In addition, large basophilic epitheliomas demonstrated an increase in both SYN and PHO activities. In contrast, most oncocytic tubules and oncocytomas characterized by abundant densely granular cytoplasm showed a reduction in the activity of G6PDH, but were intensely positive for SDH. However, a few oncocytic lesions demonstrated a decrease in both SDH and G6PDH activity. Rarely, decreased SDH and elevated G6PDH activities were observed in altered tubules resembling oncocytic tubules. It remains to be clarified whether these tubules represent a variation of the oncocytic lesions or, perhaps, another type of tubular lesion. The results indicate that basophilic and oncocytic epithelial tumors differ in their cytochemical pattern and histogenesis. In line with earlier suggestions, the basophilic tumors apparently originate from the proximal renal tubules, while the oncocytomas develop from the distal parts of the nephron. The basophilic tumors are characterized by an increased pentose phosphate pathway and glycolysis, with a corresponding reduction in mitochondrial respiration. However, the majority of the oncocytomas show an increased activity of the mitochondrial enzyme SDH, and a marked decrease in the activity of the key enzyme of the pentose phosphate pathway.     

Substrate specificity classes and the recognition signal for Salmonella type III flagellar export

Most flagellar proteins of Salmonella are exported to their assembly destination via a specialized apparatus. This apparatus is a member of the type III superfamily, which is widely used for secretion of virulence factors by pathogenic bacteria. Extensive studies have been carried out on the export of several of the flagellar proteins, most notably the hook protein (FlgE), the hook-capping protein (FlgD), and the filament protein flagellin (FliC). This has led to the concept of two export specificity classes, the rod/hook type and the filament type. However, little direct experimental evidence has been available on the export properties of the basal-body rod proteins (FlgB, FlgC, FlgF, and FlgG), the putative MS ring-rod junction protein (FliE), or the muramidase and putative rod-capping protein (FlgJ). In this study, we have measured the amounts of these proteins exported before and after hook completion. Their amounts in the culture supernatant from a flgE mutant (which is still at the hook-type specificity stage) were much higher than those from a flgK mutant (which has advanced to the filament-type specificity stage), placing them in the same class as the hook-type proteins. Overproduction of FliE, FlgB, FlgC, FlgF, FlgG, or FlgJ caused inhibition of the motility of wild-type cells and inhibition of the export of the hook-capping protein FlgD. We also examined the question of whether export and translation are linked and found that all substrates tested could be exported after protein synthesis had been blocked by spectinomycin or chloramphenicol. We conclude that the amino acid sequence of these proteins suffices to mediate their recognition and export.     

Energetics and volume changes of the intermediates in the photolysis of octopus rhodopsin at a physiological temperature

Enthalpy changes (Delta H) of the photointermediates that appear in the photolysis of octopus rhodopsin were measured at physiological temperatures by the laser-induced transient grating method. The enthalpy from the initial state, rhodopsin, to bathorhodopsin, lumirhodopsin, mesorhodopsin, transient acid metarhodopsin, and acid metarhodopsin were 146 +/- 15 kJ/mol, 122 +/- 17 kJ/mol, 38 +/- 8 kJ/mol, 12 +/- 5 kJ/mol, and 12 +/- 5 kJ/mol, respectively. These values, except for lumirhodopsin, are similar to those obtained for the cryogenically trapped intermediate species by direct calorimetric measurements. However, the Delta H of lumirhodopsin at physiological temperatures is quite different from that at low temperature. The reaction volume changes of these processes were determined by the pulsed laser-induced photoacoustic method along with the above Delta H values. Initially, in the transformation between rhodopsin and bathorhodopsin, a large volume expansion of +32 +/- 3 ml/mol was obtained. The volume changes of the subsequent reaction steps were rather small. These results are compared with the structural changes of the chromophore, peptide backbone, and water molecules within the membrane helixes reported previously.     

Ranging behavior of Mahale chimpanzees: a 16 year study

We have analyzed the ranging patterns of the Mimikire group (M group) of chimpanzees in the Mahale Mountains National Park, Tanzania. During 16 years, the chimpanzees moved over a total area of 25.2 or 27.4 km², as estimated by the grid-cell or minimum convex polygon (MCP) methods, respectively. Annually, the M group used an average of 18.4 km², or approximately 70 %, of the total home-range area. The chimpanzees had used 80 % of their total home range after 5 years and 95 % after 11 years. M group chimpanzees were observed more than half of the time in areas that composed only 15 % of their total home range. Thus, they typically moved over limited areas, visiting other parts of their range only occasionally. On average, the chimpanzees used 7.6 km² (in MCP) per month. Mean monthly range size was smallest at the end of the rainy season and largest at the end of the dry season, but there was much variability from year to year. The chimpanzees used many of the same areas every year when Saba comorensis fruits were abundant between August and January. In contrast, the chimpanzees used several different areas of their range in June. Here range overlap between years was relatively small. Over the 16 years of the study we found that the M group reduced their use of the northern part of their range and increased their frequency of visits to the eastern mountainous side of their home range. Changes in home-range size correlated positively with the number of adult females but not with the number of adult males. This finding does not support a prediction of the male-defended territory model proposed for some East African chimpanzee unit-groups.     

Genetic structure of <Emphasis Type="Italic">Cerasus jamasakura</Emphasis>, a Japanese flowering cherry,revealed by nuclear SSRs: implications for conservation

The genetic resources of a particular species of flowering cherry, Cerasus jamasakura, have high conservation priority because of its cultural, ecological and economic value in Japan. Therefore, the genetic structures of 12 natural populations of C. jamasakura were assessed using ten nuclear SSR loci. The population differentiation was relatively low (F _ST, 0.043), reflecting long-distance dispersal of seeds by animals and historical human activities. However, a neighbor-joining tree derived from the acquired data, spatial analysis of molecular variance and STRUCTURE analysis revealed that the populations could be divided into two groups: one located on Kyusyu Island and one on Honshu Island. Genetic diversity parameters such as allelic richness and gene diversity were significantly lower in the Kyushu group than the Honshu group. Furthermore, STRUCTURE analysis revealed that the two lineages were admixed in the western part of Honshu Island. Thus, although the phylogeographical structure of the species and hybridization dynamics among related species need to be evaluated in detail using several marker systems, the Kyusyu Island and Honshu Island populations should be considered as different conservation units, and the islands should be regarded as distinct seed transfer zones for C. jamasakura, especially when rapid assessments are required.     

Galectin-9 Enhances Cytokine Secretion,but Suppresses Survival and Degranulation,in Human Mast Cell Line

Galectin-9 (Gal-9), a lectin having a β-galactoside-binding domain, can induce apoptosis of Th1 cells by binding to TIM-3. In addition, Gal-9 inhibits IgE/Ag-mediated degranulation of mast cell/basophilic cell lines by binding to IgE, thus blocking IgE/Ag complex formation. However, the role of Gal-9 in mast cell function in the absence of IgE is not fully understood. Here, we found that recombinant Gal-9 directly induced phosphorylation of Erk1/2 but not p38 MAPK in a human mast cell line, HMC-1, which does not express FcεRI. Gal-9 induced apoptosis and inhibited PMA/ionomycin-mediated degranulation of HMC-1 cells. On the other hand, Gal-9 induced cytokine and/or chemokine production by HMC-1 cells, dependent on activation of ERK1/2 but not p38 MAPK. In addition, the lectin activity of Gal-9 was required for Gal-9-mediated cytokine secretion by HMC-1 cells. These observations suggest that Gal-9 has dual properties as both a regulator and an activator of mast cells.     

Exacerbation of experimental allergic asthma by augmented Th2 responses in WSX-1-deficient mice

WSX-1 (IL-27R) is a class I cytokine receptor with homology to gp130 and IL-12 receptors and is typically expressed on CD4+ T lymphocytes. Although previous reports have clarified that IL-27/WSX-1 signaling plays critical roles in both Th1 differentiation and attenuation of cell activation and proinflammatory cytokine production during some bacterial or protozoan infections, little is known about the importance of WSX-1 in cytokine-mediated diseases of allergic origin. To this aim, we took advantage of WSX-1-deficient (WSX-1(-/-)) mice and induced experimental asthma, in which Th2 cytokines are central modulators of the pathology. OVA-challenged WSX-1(-/-) mice showed marked enhancement of airway responsiveness with goblet cell hyperplasia, pulmonary eosinophil infiltration, and increased serum IgE levels compared with wild-type mice. Production of Th2 cytokines, which are largely responsible for the pathogenesis of asthma, was augmented in the lung or in the culture supernatants of peribronchial lymph node CD4+ T cells from WSX-1(-/-) mice compared with those from wild-type mice. Surprisingly, IFN-gamma production was also enhanced in WSX-1(-/-) mice, albeit at a low concentration. The cytokine overproduction, thus, seems independent from the Th1-promoting property of WSX-1. These results demonstrated that IL-27/WSX-1 also plays an important role in the down-regulation of airway hyper-reactivity and lung inflammation during the development of allergic asthma through its suppressive effect on cytokine production.     

Variation in mate recognition specificities among four Callosobruchus seed beetles

Differentiation of mate recognition systems is one of the important steps for speciation in animals. For some insects, a contact sex pheromone present on the cuticular surface is indispensable in discriminating reproductive partners. In Callosobruchus species (Coleoptera: Chrysomelidae: Bruchinae), contact sex pheromones have been found in two species, Callosobruchus chinensis (L.) and Callosobruchus maculatus (Fabricius). It was suggested, however, that these two species lacked the ability to discriminate their conspecific and/or heterosexual partners. To elucidate this inconsistency, we verified the existence of contact sex pheromones from two other species, Callosobruchus rhodesianus (Pic) and Callosobruchus analis (Fabricius). As a result, unlike C. chinensis and C. maculatus, the males of C. rhodesianus and C. analis were able to discriminate their heterosexual partners. Comparing cross‐copulation behavior, i.e., copulation behavior between two species, against these four species indicated that the mate recognition specificities were quite different. Males of C. rhodesianus and C. analis had highly species‐specific mating behavior, whereas males of C. chinensis and C. maculatus were much less specific. These results indicate that variation in mate recognition can arise even among congeneric species living in a sympatric environment, and this variation might have arisen during species differentiation. Based on our results in combination with previous reports on interspecific competition, we suggest that the observed asymmetric cross‐copulation behavior might be, at least partially, an adaptation for surviving interspecific competition.     

Genetically engineered rice containing larger amounts of nicotianamine to enhance the antihypertensive effect

Nicotianamine (NA), a metal chelator ubiquitous in higher plants, serves as an antihypertensive substance in humans. To engineer a novel antihypertensive rice that contains larger amounts of NA, the barley NA synthase gene, HvNAS1 , was introduced into rice via Agrobacterium -mediated transformation. The introduced HvNAS1 was driven by pGluB-1 , which induces strong gene expression in the endosperm of rice seeds. The NA content in transgenic rice seeds was up to fourfold greater than that in non-transgenic rice seeds. The Cre/ loxP DNA excision (CLX) system was used to remove the selectable marker gene for antibiotic resistance. Furthermore, the transgenic rice was crossed with a cleistogamous mutant to prevent gene transfer via pollen dispersal. These two modifications may minimize public concern with regard to the use of this transgenic rice.