bluuebunny/crossref_metadata_2025 huggingface dataset:
PP2C family members play key roles in regulation of cell survival and apoptosis
Authors: Tamura Shinri , Toriumi Shinnosuke , Saito Jun‐ichi , Awano Kenjiro , Kudo Tada‐aki , Kobayashi Takayasu
Published: April 2006

Although unlimited proliferation of cancer cells is supported by multiple signaling pathways involved in the regulation of proliferation, survival, and apoptosis, the molecular mechanisms coordinating these different pathways to promote the proliferation and survival of cancer cells have remained unclear. SAPK and integrin‐ILK signaling pathways play key roles in the promotion of apoptosis and cell proliferation/survival, respectively. Studies of TNFα‐ and H2O2‐induced apoptosis revealed that ASK1, a component of the SAPK system, mediates the TNFα and H2O2signaling of apoptosis. ASK1 is activated by autophosphorylation of a specific threonine residue (T845) following TNFα stimulation. Our recent studies indicate that PP2Cɛ, a member of the PP2C family, associates with and inactivates ASK1 by dephosphorylating T845. In contrast, PP2Cδ/ILKAP, a second PP2C family member, activates ASK1 by enhancing cellular phosphorylation of T845. PP2Cδ/ILKAP also forms a complex with ILK1 to inhibit the GSK3β‐mediated integrin‐ILK1 signalingin vivo, inhibiting cell cycle progression. These observations raise the possibility that PP2Cδ/ILKAP acts to control the cross‐talk between integrin‐induced and TNFα‐induced signaling pathways, inhibiting the former and stimulating the latter, thereby inhibiting proliferation and survival and promoting the apoptosis of cancer cells. (Cancer Sci2006; 97: 563–567)
Endogenous markers of hypoxia/anaerobic metabolism and anemia in primary colorectal cancer
Authors: Koukourakis Michael I. , Giatromanolaki Alexandra , Polychronidis Alexandros , Simopoulos Costantinos , Gatter Kevin C. , Harris Adrian L. , Sivridis Efthimios
Published: April 2006

Anemia has been implicated in the decreased oxygen tension noted within the tumor environment. In a series of 79 colorectal adenocarcinomas we investigated the role of anemia in activating molecular pathways regulated by hypoxia. Preoperative Hb levels were correlated with the immunohistochemical expression of HIF1α and HIF2α, LDH5, GLUT1, VEGF, DEC1 and BNIP3, and with angiogenesis and the cancer cell proliferation index. Upregulation of HIF1α and HIF2α proteins, found in 43% and 44.3% of cases, respectively, was not related to anemia (Hb < 10 g%). This is in agreement with other studies suggesting that HIF activation occurs for various reasons, such as poor or irregular vascularity, or oncogene activation. Nevertheless, low Hb levels (<10 g%) were linked to activated anaerobic metabolism (LDH5 overexpression) in a subset of tumors not expressing HIF1α (P < 0.01). Overexpression of HIFs, whether linked to anemia or not, was associated with a number of factors related to tumor aggressiveness (assessed as local invasion and nodal metastasis), anaerobic metabolism and intratumoral acidosis (LDH5, GLUT1; increased glucose metabolism to lactate), activation of genes related to necrosis (BNIP3) and angiogenesis (VEGF). Expression of BNIP3 emerged as the strongest independent factor related to transmural invasion and metastasis to lymph nodes. Identification of specific patterns of the hypoxia molecular cascade activated in cancer cells might help in developing specific therapeutic policies. (Cancer Sci2006; 97: 582–588)
Chemotherapeutic efficacy of paclitaxel in combination with Withania somnifera on benzo(a)pyrene‐induced experimental lung cancer
Authors: Senthilnathan Palaniyandi , Padmavathi Radhakrishnan , Magesh Venkatraman , Sakthisekaran Dhanapal
Published: April 2006

Lung cancer is one of the leading causes of cancer death in the world and is notoriously difficult to treat effectively. In the present study, male Swiss albino mice were divided into five groups of six animals each: group I animals received corn oil orally and served as a control; group II cancer‐induced animals received benzo(a)pyrene (50 mg/kg bodyweight dissolved in corn oil, orally) twice weekly for four successive weeks; group III cancer‐bearing animals (after 12 weeks of induction) were treated with paclitaxel (33 mg/kg bodyweight, i.p.) once weekly for 4 weeks; group IV cancer‐bearing animals were treated with paclitaxel along with Withania somnifera (400 mg/kg bodyweight) orally once weekly for 4 weeks; and group V animals constituted the drug control treated with paclitaxel along with W. somnifera. The serum, lung and liver were investigated biochemically for aryl hydrocarbon hydroxylase, γ‐glutamyl transpeptidase, 5′‐nucleotidase, lactate dehydrogenase and protein‐bound carbohydrate components (hexose, hexosamine and sialic acid). These enzyme activities were increased significantly in cancer‐bearing animals compared with control animals. The elevation of these in cancer‐bearing animals was indicative of the persistent deteriorating effect of benzo(a)pyrene in cancer‐bearing animals. Our data suggest that paclitaxel, administered with W. somnifera, may extend its chemotherapeutic effect through modulating protein‐bound carbohydrate levels and marker enzymes, as they are indicators of cancer. The combination of paclitaxel with W. somnifera could effectively treat the benzo(a)pyrene‐induced lung cancer in mice by offering protection from reactive oxygen species damage and also by suppressing cell proliferation. (Cancer Sci 2006; 97: 658–664)
Circadian gene mPer2 overexpression induces cancer cell apoptosis
Authors: Hua Hui , Wang Yueqi , Wan Chaomin , Liu Yanyou , Zhu Bin , Yang Chunlei , Wang Xiaojia , Wang Zhengrong , Cornelissen–Guillaume Germaine , Halberg Franz
Published: April 2006

The Period2 gene, an indispensable component of the circadian clock, not only modulates circadian oscillations, but also regulates organic function. We examined whether overexpression of the mouse Period2 gene (mPer2) in tumor cells influences cell growth and induces apoptosis. Overexpression of PERIOD2 in the mouse Lewis lung carcinoma cell line (LLC) and mammary carcinoma cell line (EMT6) results in reduced cellular proliferation and rapid apoptosis, but not in NIH 3T3 cells. Overexpressed mPER2 also altered the expression of apoptosis‐related genes. The mRNA and protein levels of c‐Myc, Bcl‐XL and Bcl‐2 were downregulated, whereas the expression of p53 and bax was upregulated in mPER2‐overexpressing LLC cells compared with control cells transferred with empty plasmid. Our results suggest that the circadian gene mPeriod2 may play an important role in tumor suppression by inducing apoptotic cell death, which is attributable to enhanced pro‐apoptotis signaling and attenuated anti‐apoptosis processes. (Cancer Sci 2006; 97: 589–596)
PARK2/PACRG polymorphisms and susceptibility to typhoid and paratyphoid fever
Authors: Ali S , Vollaard A M , Widjaja S , Surjadi C , Van De Vosse E , Van Dissel J T
Published: April 2006

Host genetic factors may contribute to susceptibility to and outcome in infectious diseases. Recently polymorphisms in PARK2/PACRG, a gene cluster linked to ubiquitination and proteasome-mediated protein degradation, were found to be associated with manifest infection by M. leprae. Here, we address whether these polymorphisms are associated with susceptibility to infection with Salmonella typhi and S. paratyphi A, intracellular pathogens that upon infection of humans share with mycobacteria aspects of the hosts’ immune response. The polymorphisms of PARK_e01(−697), PARK2_e01(−2599), rs1333955 and rs1040079 were analysed by polymerase chain reaction and restriction fragment length polymorphism in a case-control study of typhoid and paratyphoid fever patients in an endemic area in Jakarta, Indonesia. For this study, samples were obtained from patients with blood culture-confirmed typhoid fever (n = 90), paratyphoid fever (n = 26) and fever controls (n = 337) in a passive, community-based surveillance and compared to those of randomly selected community controls (n = 322) from the same city area. The PARK2_e01(−2599) allele T was significantly associated with typhoid and paratyphoid fever (OR: 1·51, 95%CI: 1·02–2·23) but the other polymorphisms, PARK2_e01(−697), rs1333955 and rs1040079, were not associated. Although within the PARK2/PACRG gene cluster the PARK2_e01(−2599) allele T was most strongly associated with leprosy (OR∼ 3–5), the association with typhoid is much less strong. Our findings suggest that this polymorphism in PARK2/PACRG plays a small but significant role in susceptibility to the intracellular pathogens S. typhi and S. paratyphi.
Preferential recognition of the phosphorylated major linear B-cell epitope of La/SSB 349–368aa by anti-La/SSB autoantibodies from patients with systemic autoimmune diseases
Authors: Terzoglou A G , Routsias J G , Avrameas S , Moutsopoulos H M , Tzioufas A G
Published: April 2006

Sera from patients with primary Sjögren Syndrome (pSS) or Systemic Lupus Erythematosus (SLE) often contain autoantibodies directed against La/SSB. The sequence 349–368aa represents the major B-cell epitope of La/SSB, also it contains, at position 366, a serine aminoacid residue which constitutes the main phosphorylation site of the protein. In this study we investigated the differential recognition of the 349–368aa epitope and its phosphorylated form by antibodies found in sera from patients with systemic autoimmune diseases. Peptides corresponding to the sequence of the unphosphorylated (pep349–368aa) and the phosphorylated form (pep349–368aaPh) of the La/SSB epitope 349–368aa, as well as to a truncated form spanning the sequence 349–364aa and lacking the phosphorylation site (pep349–364aa), were synthesized. Sera from 53 patients with pSS and SLE with anti-La/SSB specificity, 30 patients with pSS and SLE without anti-La/SSB antibodies, 25 patients with rheumatoid arthritis and 32 healthy individuals were investigated by ELISA experiments. Autoantibodies to pep349–368aaPh were detected in sera of anti-La/SSB positive patients with a higher prevalence compared to the pep349–368aa (66%versus 45%). Pep349–368aaPh inhibited the antibody binding almost completely (92%), while pep349–368aa inhibited the binding only partially (45%). Anti-La/SSB antibodies presented a higher relative avidity for the phosphorylated than the unphosphorylated peptide. Immunoadsorbent experiments using the truncated peptide pep349–364aa indicated that the flowthrough showed a selective specificity for pep349–368aaPh, while the eluted antibodies reacted with both peptide analogues of the La/SSB epitope. These data suggest that sera from pSS and SLE patients with anti-La/SSB reactivity possess autoantibodies that bind more frequently and with a higher avidity to the phosphorylated major B-cell epitope of the molecule.
Effect of investigator disturbance in experimental forensic entomology: succession and community composition
Authors: De Jong G. D. , Hoback W. W.
Published: April 2006

Abstract. Carrion insect succession studies have historically used repeated sampling of single or a few carcasses to produce data, either weighing the carcasses, removing a qualitative subsample of the fauna present, or both, on every visit over the course of decomposition and succession. This study, conducted in a set of related experimental hypotheses with two trials in a single season, investigated the effect that repeated sampling has on insect succession, determined by the number of taxa collected on each visit and by community composition. Each trial lasted at least 21 days, with daily visits on the first 14 days. Rat carcasses used in this study were all placed in the field on the same day, but then either sampled qualitatively on every visit (similar to most succession studies) or ignored until a given day of succession, when they were sampled qualitatively (a subsample) and then destructively sampled in their entirety. Carcasses sampled on every visit were in two groups: those from which only a sample of the fauna was taken and those from which a sample of fauna was taken and the carcass was weighed for biomass determination. Of the carcasses visited only once, the number of taxa in subsamples was compared to the actual number of taxa present when the carcass was destructively sampled to determine if the subsamples adequately represented the total carcass fauna. Data from the qualitative subsamples of those carcasses visited only once were also compared to data collected from carcasses that were sampled on every visit to investigate the effect of the repeated sampling. A total of 39 taxa were collected from carcasses during the study and the component taxa are discussed individually in relation to their role in succession. Number of taxa differed on only one visit between the qualitative subsamples and the actual number of taxa present, primarily because the organisms missed by the qualitative sampling were cryptic (hidden deep within body cavities) or rare (only represented by very few specimens). There were no differences discovered between number of taxa in qualitative subsamples from carcasses sampled repeatedly (with or without biomass determinations) and those sampled only a single time. Community composition differed considerably in later stages of decomposition, with disparate communities due primarily to small numbers of rare taxa. These results indicate that the methods used historically for community composition determination in experimental forensic entomology are generally adequate.
Characterization of a conjugative transposon integrase, IntDOT
Authors: Malanowska Karolina , Salyers Abigail A. , Gardner Jeffrey F.
Published: April 2006

Sequence analysis revealed that the integrase of the Bacteroides conjugative transposon CTnDOT (IntDOT) might be a member of the tyrosine recombinase family because IntDOT has five of six highly conserved residues found in the catalytic domains of tyrosine recombinases. Yet, IntDOT catalyses a reaction that appears to differ in some respects from well‐studied tyrosine recombinases such as that of phage λ. To assess the importance of the conserved residues, we changed residues in IntDOT that align with conserved residues in tyrosine recombinases. Some substitutions resulted in a complete loss or significant decrease of integration activity in vivo. The ability of the mutant proteins to cleave and ligate CTnDOT attachment site (attDOT) DNA in vitro in general paralleled the in vivo results, but the H345A mutant, which had a wild‐type level of integration in vivo, exhibited a slightly lower level of cleavage and ligation in vitro. Our results confirm the hypothesis that IntDOT belongs to the tyrosine recombinase family, but the catalytic core of the protein seems to have somewhat different organization. Previous DNA sequence analyses showed that CTnDOT att sites contain 5 bp non‐homologous coupling sequences which were assumed to define the putative staggered sites of cleavage. However, cleavage assays showed that one of the cleavage sites is 2 bp away from the junction of CTnDOT and coupling sequence DNA. The site is in a region of homology that is conserved in CTnDOT att sites.
The Campylobacter jejuni PEB1a adhesin is an aspartate/glutamate‐binding protein of an ABC transporter essential for microaerobic growth on dicarboxylic amino acids
Authors: Del Rocio Leon‐Kempis Maria , Guccione Edward , Mulholland Francis , Williamson Michael P. , Kelly David J.
Published: April 2006

The PEB1a protein of the gastrointestinal pathogen Campylobacter jejuni mediates interactions with epithelial cells and is an important factor in host colonization. Cell fractionation and immunoblotting showed that PEB1a is most abundant in the periplasm of C. jejuni, and is detectable in the culture supernatant but not in the inner or outer membrane. The protein is homologous with periplasmic‐binding proteins associated with ABC transporters and we show by fluorescence spectroscopy that purified recombinant PEB1a binds l‐aspartate and l‐glutamate with sub µM Kd values. Binding of l‐14C‐aspartate or l‐14C‐glutamate was strongly out‐competed by excess unlabelled aspartate or glutamate but only poorly by asparagine and glutamine. A mutant in the Cj0921c gene, encoding PEB1a, was completely unable to transport 5 µM l‐14C‐glutamate and showed a large reduction (∼20‐fold) in the rate of l‐14C‐aspartate transport compared with the wild type. Although microaerobic growth of this mutant was little affected in complex media, growth on aspartate or glutamate in defined media was completely prevented, whereas growth with serine was similar to wild type. 1H‐NMR analysis of the culture supernatants of the Cj0921c mutant showed some utilization of aspartate but not glutamate, consistent with the transport data. It is concluded that in addition to the established role of PEB1a as an adhesin, the PEB1 transport system plays a key role in the utilization of aspartate and glutamate, which may be important in vivo carbon sources for this pathogen.
Regulated expression of glutamyl‐tRNA synthetase is directed by a mobile genetic element in the cyanobacterium Tolypothrix sp. PCC 7601
Authors: Luque Ignacio , Andújar Alfonso , Jia Lin , Zabulon Gérald , De Marsac Nicole Tandeau , Flores Enrique , Houmard Jean
Published: April 2006

The genome of Tolypothrix sp. PCC 7601 carries two copies of a novel insertion sequence, ISTosp1. One of the two copies is located upstream of the gene encoding glutamyl‐tRNA synthetase, an enzyme playing a key role in protein and pigment synthesis. The tnpA gene of the IS element and gltX were co‐transcribed and their expression was transiently upregulated upon retrieval of the ammonium source irrespective of whether nitrate or no nitrogen source were available. The second copy is also transcribed and shows a similar regulatory pattern. Structural elements of the promoter (−10 and −35 sequences) directing the expression of the tnpA‐gltX operon have been localized within the IS. Regulatory sequences involving the NtcA transcription factor in the control of tnpA‐gltX expression were found both within and in sequences upstream of the insertion element. The expression of gltX in a closely related cyanobacterium, Nostoc sp. PCC 7120, which lacks the insertion upstream of gltX, decreased upon ammonium retrieval, a regulatory pattern that markedly differs from that observed in Tolypothrix sp. PCC 7601. ISTosp1 constitutes a good example of how cells can make use of a transposable element to evolve an original regulatory mechanism.
M. tuberculosis Rv2252 encodes a diacylglycerol kinase involved in the biosynthesis of phosphatidylinositol mannosides (PIMs)
Authors: Owens Róisín M. , Hsu Fong Fu , VanderVen Brian C. , Purdy Georgiana E. , Hesteande Elizabeth , Giannakas Peter , Sacchettini James C. , McKinney John D. , Hill Preston J. , Belisle John T. , Butcher Barbara A. , Pethe Kevin , Russell David G.
Published: April 2006

Phosphorylated lipids play important roles in biological systems, not only as structural moieties but also as modulators of cellular function. Phospholipids of pathogenic bacteria are known to play roles both as membrane components and as factors that modulate the infectious process. Mycobacterium tuberculosis is, however, noteworthy in that it has an extremely diverse repertoire of biologically active phosphorylated lipids that, in the absence of a specialized protein translocation system, appear to constitute the main means of communication with the host. Many of these lipids are derived from phosphatidylinositol (PI) that is differentially processed to give rise to phosphatidylinositol mannosides (PIMs) or lipoarabinomannan. In preliminary studies on the lipid processing enzymes associated with the bacterial cell wall, a kinase activity was noted that gave rise to a novel lipid species released by the bacterium. It was determined that this kinase activity was encoded by the ORF Rv2252. Rv2252 demonstrates the capacity to phosphorylate various amphipathic lipids of host and bacterial origin, in particular a M. tuberculosis derived diacylglycerol. Targeted deletion of the rv2252 gene resulted in disruption of the production of certain higher order PIM species, suggesting a role for Rv2252 in the biosynthetic pathway of PI, a PIM precursor.
Effects of serotonin transporter inhibition on gastrointestinal motility and colonic sensitivity in the mouse
Authors: Coates M. D. , Johnson A. C. , Greenwood‐van Meerveld B. , Mawe G. M.
Published: April 2006

Abstract  Serotonin‐selective reuptake transporter (SERT) expression is decreased in animal models of intestinal inflammation and in individuals with inflammatory bowel disease (IBD) or irritable bowel syndrome (IBS), and it is possible that resultant changes in intestinal serotonin signalling contribute to the manifestation of clinical features associated with these disorders. The objective of this investigation was to determine whether inhibition of SERT function leads to changes in gut motility and sensitivity. Mice underwent a 14‐day treatment with the SERT inhibitor, paroxetine (20 mg kg−1), or vehicle (saline/propylene glycol). Gastrointestinal (GI) transit following charcoal gavage, colonic motility, stool frequency and visceromotor responses to colorectal distension were evaluated. In mice treated with paroxetine, stool output was decreased, upper GI transit was delayed, and colonic sensitivity to a nociceptive stimulus was attenuated. These results demonstrate that reduced SERT function (via pharmacological blockade) significantly alters GI motility and sensitivity in mice, and support the concept that altered SERT expression and function could contribute to symptoms associated with IBS and IBD.
Patterns of SPARC expression and basement membrane intactness at the tumour–brain border of invasive meningiomas
Authors: Schittenhelm J. , Mittelbronn M. , Roser F. , Tatagiba M. , Mawrin C. , Bornemann A.
Published: April 2006

The matricellular glycoprotein SPARC (secreted protein, acidic and rich in cysteine), also termed osteonectin, has been found to regulate the invasive behaviour of several tumour types by interacting with basement membrane constituents. Brain invasive meningiomas are supposed to disrupt the pial‐glial basement membrane. In the present study we aimed at determining the relationship of basement membrane intactness and SPARC protein expression at the meningioma–brain border. Sections of 51 brain‐invasive meningiomas (31 meningothelial meningiomas WHO grade I, 11 atypical WHO grade II, and nine anaplastic WHO grade III tumours) were immunolabelled with antibodies against SPARC, epithelial membrane antigen (EMA), collagen IV and glial fibrillary acidic protein (GFAP). Twenty‐two non‐invasive WHO grade I meningothelial meningiomas were included in the study for comparison. At the tumour–brain border of invasive meningiomas, spindle‐shaped tumour cells expressed SPARC. The number of tumours containing SPARC+ spindle cells did not differ significantly between WHO grades. By contrast, the number of WHO grade I tumours expressing collagen IV (15/31) was highly significantly elevated when compared with WHO grade II (1/11) and WHO grade III (0/9) (both P < 0.0001). There was an inverse relationship of the presence of SPARC+ spindle cells and basement membrane material. In conclusion, the destruction of the basement membrane is correlated with meningioma malignancy grade whereas the expression of SPARC protein at the tumour–brain border is not. Destruction of the basement membrane and appearance of SPARC+ spindle cells are not coincident during the course of brain invasion by meningiomas.
Tonoplast‐located GmCLC1 and GmNHX1 from soybean enhance NaCl tolerance in transgenic bright yellow (BY)‐2 cells
Authors: LI WING‐YEN FRANCISCA , WONG FUK‐LING , TSAI SAU‐NA , PHANG TSUI‐HUNG , SHAO GUIHUA , LAM HON‐MING
Published: April 2006

Genes encoding ion transporters that regulate ion homeostasis in soybean have not been carefully investigated. Using degenerate primers, we cloned a putative chloride channel gene (GmCLC1) and a putative Na+/H+ antiporter gene (GmNHX1) from soybean. Confocal microscopic studies using yellow fluorescent fusion proteins revealed that GmCLC1 and GmNHX1 were both localized on tonoplast. The expressions of GmCLC1 and GmNHX1 were both induced by NaCl or dehydration stress imposed by polyethylene glycol (PEG). Using mitochondrial integrity and cell death as the damage indicators, a clear alleviation under NaCl stress (but not PEG stress) was observed in both GmCLC1 and GmNHX1 transgenic cells. Using fluorescent dye staining and quenching, respectively, a higher concentration of chloride ion (Cl–) or sodium ion (Na+) was observed in isolated vacuoles in the cells of GmCLC1 and of GmNHX1 transgenic lines. Our result suggested that these vacuolar‐located ion transporters function to sequester ions from cytoplasm into vacuole to reduce its toxic effects.
Genetic diversity and host differentiation among isolates of Phytophthora infestans from cultivated potato and wild solanaceous hosts in Peru
Authors: Garry G. , Forbes G. A. , Salas A. , Santa Cruz M. , Perez W. G. , Nelson R. J.
Published: April 2006

To test the hypothesis that isolates of Phytophthora infestans attacking wild Solanaceae exhibit specialization for particular host species, 115 isolates of P. infestans were collected from cultivated potatoes, nontuber‐bearing Solanum spp. of the Basarthrum section and wild tomatoes from five departments in the northern and central highlands of Peru, and characterized using several neutral markers. All isolates belonged to one of four clonal lineages described previously in Peru: EC‐1, US‐1, PE‐3 and PE‐7. There was a strong association of three lineages with host species: PE‐3 was only isolated from cultivated potato, while PE‐7 and US‐1 were only isolated from nontuber‐bearing Solanum spp. (Basarthrum section and wild tomatoes). EC‐1 was isolated from all host groups sampled. A subset (n = 74) of the isolates was evaluated for metalaxyl resistance. High levels of resistance were found almost exclusively in EC‐1 and PE‐3, while US‐1 and PE‐7 isolates were generally sensitive. In a detached‐leaf assay for lesion diameter using five EC‐1 isolates from S. caripense and seven EC‐1 isolates from cultivated potato, there was a significant interaction between isolate origin and inoculated host, caused by higher aggressiveness of EC‐1 from cultivated potato on its host of origin. In a comparison of EC‐1 (seven isolates from cultivated potato) and US‐1 (three isolates from S. caripense), each pathogen lineage was more aggressive on its original host species, causing a highly significant interaction between isolate origin and inoculated host. Wild tomatoes and nontuber‐bearing Solanum spp. harbour several pathogen lineages in Peru and could serve as reservoirs of inoculum that might contribute to epidemics on potato or tomato. Potential risks associated with the use of wild Solanum hosts as sources of resistance to P. infestans are discussed.
AtNAP, a NAC family transcription factor, has an important role in leaf senescence
Authors: Guo Yongfeng , Gan Susheng
Published: April 2006

Leaf senescence is a unique developmental process that is characterized by massive programmed cell death and nutrient recycling. The underlying molecular regulatory mechanisms are not well understood. Here we report the functional analysis of AtNAP, a gene encoding a NAC family transcription factor. Expression of this gene is closely associated with the senescence process of Arabidopsis rosette leaves. Leaf senescence in two T‐DNA insertion lines of this gene is significantly delayed. The T‐DNA knockout plants are otherwise normal. The mutant phenotype can be restored to wild‐type by the intact AtNAP, as well as by its homologs in rice and kidney bean plants that are also upregulated during leaf senescence. Furthermore, inducible overexpression of AtNAP causes precocious senescence. These data strongly suggest that AtNAP and its homologs play an important role in leaf senescence in Arabidopsis and possibly in other plant species.
TT8 controls its own expression in a feedback regulation involving TTG1 and homologous MYB and bHLH factors, allowing a strong and cell‐specific accumulation of flavonoids in Arabidopsis thaliana
Authors: Baudry Antoine , Caboche Michel , Lepiniec Loïc
Published: April 2006

The control of TT8 expression was investigated in this study, and it was demonstrated that it constitutes a major regulatory step in the specific activation of the expression of flavonoid structural genes. First, the GUS activity generated in planta from a TT8::uidA construct revealed cell‐specific activation of the TT8 promoter consistent with the known involvement of the TT8 bHLH factor in proanthocyanidin, anthocyanin and mucilage biosynthesis. Moreover, the activity of this reporter construct was strongly affected in ttg1, TT2 overexpressers (OE), and PAP1‐OE, suggesting interplay between TT2, PAP1, TTG1 and the activation of the TT8 promoter in planta. To further investigate the mechanisms involved, we used 35S::TT2‐GR and 35S::TTG1‐GR transgenic plants (expressing fusion proteins with the glucocorticoid receptor), as well as one‐hybrid experiments, to determine the direct effect of these factors on TT8 expression. Interestingly, in vivo binding of TT2 and PAP1 to the TT8 promoter was dependent on the simultaneous expression of TT8 or the homologous bHLH factors GL3 and EGL3. Consistent with these results, the activity of the TT8::uidA reporter was strongly affected in the seed endothelium of a tt8 mutant. Similarly, a strong decrease in the level of TT8 mRNA was detected in the siliques of a gl3 × egl3 mutant and in plants that express a dominant negative form of the PAP1 protein, suggesting that TT8 expression is controlled by different combinations of MYB and bHLH factors in planta. The importance of this positive feedback mechanism in the strong and specific induction of proanthocyanidin biosynthesis in the seed coat of Arabidopsis thaliana is discussed.
The Arabidopsis SPA1 gene is required for circadian clock function and photoperiodic flowering
Authors: Ishikawa Masaki , Kiba Takatoshi , Chua Nam‐Hai
Published: April 2006

Arabidopsis phytochrome A (phyA) regulates not only seed germination and seedling de‐etiolation but also circadian rhythms and flowering time in adult plants. The SUPPRESSOR OF PHYA‐105 (SPA1) acts as a negative regulator of phyA‐mediated de‐etiolation of young seedlings, but its roles in adult plants have not yet been described. Here, we show that SPA1 is involved in regulating circadian rhythms and flowering time in plants. Under constant light, the abundance of SPA1 protein exhibited circadian regulation, whereas under constant darkness, SPA1 protein levels remained unchanged. These results indicate that the SPA1 protein is controlled by the circadian clock and light signals. In addition, the spa1‐3 mutation slightly shortened the circadian period of CCA1, TOC1/PRR1 and SPA1 transcript accumulation under constant light. Phenotypic analysis showed that the spa1‐3 mutant flowers early under short‐day (SD) but not long‐day (LD) conditions. Consistent with this finding, transcripts encoding flowering locus T (FT), which promotes flowering, increased in spa1‐3 under only SD conditions, although the CONSTANS (CO) transcript level was not affected under either SD nor LD conditions. Our results indicate that SPA1 not only negatively controls phyA‐mediated signaling in seedlings, but also regulates circadian rhythms and flowering time in plants.
Prevalence of celiac disease among school children in Punjab, North India
Authors: Sood Ajit , Midha Vandana , Sood Neena , Avasthi Gurcharan , Sehgal Alok
Published: April 2006

Background:  Celiac disease, as of today, is said to exist in almost all parts of the world, although it is rare among people of purely African–Caribbean, Japanese and Chinese background. The disease has also been considered uncommon in India until recently. Hospital records have revealed an increasing trend of the disease in predominantly wheat‐eating areas of North India. The aim of the present study was to determine the prevalence of celiac disease among school children in Punjab, North India.

Methods:  The study was carried out in the Ludhiana district of Punjab, Northern India. A total of 4347 children aged 3–17 years attending different schools were enrolled. A structured questionnaire was used to collect sociodemographic data and symptoms and signs related to celiac disease and various sociodemographic factors. The screening for celiac disease for the suspected celiacs was done by testing for antitissue transglutaminase (anti‐tTG) by indirect solid‐phase immunometric assay (ELISA). All children with high anti‐tTG whose parents consented underwent upper gastrointestinal endoscopy for small bowel biopsy from the second part of the duodenum. Histopathology was expressed according to the Marsh classification of 1992. Follow up was carried out among children who were put on a gluten‐restricted diet, at monthly intervals for 3 months and every 3 months thereafter. The diagnosis of celiac disease was established on the basis of the revised European Society of Paediatric Gastroenterologists and Nutritionists (ESPGAN) criteria (confirmed cases).

Results:  A total of 4347 school children (1967 girls, 2380 boys, age range 3–17 years) were screened for celiac disease. Out of these, 198 suspected children were identified for further evaluation. Twenty‐one children tested positive for anti‐tTG assay (10.6%, 95% confidence interval: 16.91–34.79). Seventeen of these 21 children agreed to undergo biopsy; of these, 14 had histological changes consistent with celiac disease and all these 14 children had clinical response to gluten restriction. Three children with high anti‐tTG had normal mucosa on duodenal biopsy and were not labelled as being in the celiac disease group. In the final analysis the disease prevalence was one in 310 children.

Conclusions:  This is the first study on celiac disease prevalence among school children from India. Although this disease frequency of one in 310 is thought to be an under‐assessment, it clearly shows that celiac disease is not rare in wheat‐eating areas of North India.
Difference in localization of esophageal mucosal breaks among grades of esophagitis
Authors: Katsube Tomoko , Adachi Kyoichi , Furuta Kenji , Miki Masaharu , Fujisawa Tomoo , Azumi Takane , Kushiyama Yoshinori , Kazumori Hideaki , Ishihara Shunji , Amano Yuji , Kinoshita Yoshikazu
Published: April 2006

Background:  Gastroesophageal reflux occurs mainly during the daytime in patients with Los Angeles grade A esophagitis, but predominantly during the night in patients with grade C and D esophagitis. The purpose of the present paper was to investigate whether this difference in the pattern of gastroesophageal reflux influences the circumferential localization of erosions in the esophageal wall.

Methods:  The subjects were 394 consecutive patients diagnosed endoscopically as having reflux esophagitis (grade A, n = 223; B, n = 93; C, n = 53; D, n = 25 cases). Their endoscopic films were reviewed retrospectively to determine the circumferential location of esophageal mucosal breaks, and also the prevalence and size of hiatal hernia (HH).

Results:  The numbers of mucosal breaks analyzed in patients with grade A, B and C esophagitis were 321, 173 and 54, respectively. Patients with grade A and B esophagitis had longitudinal mucosal breaks mainly in the right‐anterior wall of the lower esophagus, whereas patients with grade C esophagitis had transverse mucosal breaks mainly in the posterior wall. The prevalence and size of HH was significantly higher and larger, respectively, in patients with grade C or D esophagitis than in those with grade A and B esophagitis.

Conclusion:  The circumferential location of esophageal mucosal breaks differs significantly among different grades of esophagitis.
Treatment effects and predictors of a 24‐week course of interferon α‐2b plus ribavirin combination therapy for patients with chronic hepatitis C
Authors: Tomimatsu Masahiko , Aizawa Yoshio , Chuganji Yoshimichi , Ishizuka Hideo , Fujita Yoshiyuki , Aizawa Ryouichi , Abe Hiroshi , Matsuda Takako , Itou Yumi , Nakanishi Hiroyuki , Ushiyama Hisashi , Higuchi Teruhisa , Fujimoto Tomoko , Endou Hitoshi , Iga Daijiro , Ohta Kazuki , Kuroda Hiroyuki
Published: April 2006

Background and Aims:  In chronic hepatitis C patients with genotype 1b and a high viral load, the sustained virological response (SVR) rate remained as low as 2‐3% with conventional interferon (IFN) monotherapy, but improved to more than 20% with IFN α‐2b plus ribavirin combination therapy. This study examined the therapeutic effects and predictors of this combination therapy.

Methods:  Subjects were 105 patients with chronic hepatitis C (73 males, 32 females) with a median age of 53 years (range 19–70 years). Seventy‐two patients had genotype lb and 33 patients had genotype 2 (2a or 2b). Six million units (MU) or 10 MU of IFN α‐2b was administered by intramuscular injection six times a week for the first 2 weeks, and the same amount of IFN was administered three times a week for the following 22 weeks. During the IFN administration period, 600–800 mg of oral ribavirin was administered daily. Patients who were hepatitis C virus (HCV)‐RNA negative 24 weeks after the completion of administration were defined as SVR.

Results:  The overall SVR rate was 39%; 22.2% for the genotype 1b group and 75.8% for the genotype 2 group, and the difference between the groups was significant (P < 0.0001). Multivariate logistic regression analysis indicated that the factors that contributed to SVR include genotype 2, age (younger than 53 years), and an increase in Th2 measured by flow cytometry before and 4 weeks after start of treatment.

Conclusions:  The overall SVR rate of IFN α‐2b plus ribavirin combination therapy for 24 weeks was 39%, and contributing factors for SVR rate include genotype 2, age younger than 53 years and elevated Th2.
Clinical significance of measuring blood coagulation factor XIIIA regularly and continuously in patients with Crohn’s disease
Authors: Higaki Shingo , Nakano Kaori , Onaka Sachiko , Amano Ayaka , Tanioka Yukari , Harada Katsunori , Hashimoto Shinichi , Sakaida Isao , Okita Kiwamu
Published: April 2006

Background:  The aim of the present paper was to determine the mechanism by which the level of coagulation factor XIIIA declines during the active phase of Crohn’s disease.

Methods:  The relationship between the activity of Crohn’s disease and factor XIIIA was observed in 31 patients with Crohn’s disease prospectively. The relationship between factor XIIIA and thrombin–antithrombin III complex (TAT), factor XIIIA expressed on the surface of peripheral monocytes, was also evaluated.

Results:  During the first year, there were 11 patients with Crohn’s disease in the active phase and 20 patients who remained in remission. The average of the lowest level of factor XIIIA among the patients in the active phase was 60.1%. The average of the lowest level of factor XIIIA in patients who remained in remission was 78.1% (P = 0.049). Among the 20 patients who remained in remission for 1 year, eight patients had factor XIIIA levels below 70%. Six of them required surgical enterectomies, on average 2 years and 1 month later. However, none of the 12 patients who remained in remission for 1 year and who never had factor XIIIA levels <70% had a surgical enterectomy during follow up of 4 years and 6 months (P = 0.002). The decline of factor XIIIA was not due to increased consumption secondary to blood coagulation (TAT), nor was it due to a decline in the function of monocytes that produce factor XIIIA.

Conclusion:  Factor XIIIA declines during the active phase of Crohn’s disease because it might be consumed in the repair of injured tissue.
Inhibitory effects of Ligusticum chuanxiong on the proliferation of rat hepatic stellate cells
Authors: Lin Yun‐Lian , Lee Ting‐Fang , Huang Yeh‐Jeng , Huang Yi‐Tsau
Published: April 2006

Background and Aims:  Platelet‐derived growth factor (PDGF) is a very potent mitogen for hepatic stellate cells (HSC) in hepatic fibrogenesis. Ligusticum chuanxiong Hort. (LC), a traditional Chinese herb used for cerebrovascular diseases, has been shown to exert anti‐inflammatory and free radical scavenging effects. The aims of the present study were to investigate the effects of LC extract on the proliferation‐related biomarkers in a rat HSC cell line (HSC‐T6) stimulated with PDGF.

Methods:  DNA synthesis via bromodeoxyuridine (BrdU) incorporation, cell cycle related proteins and apoptosis markers were determined to evaluate the inhibitory effects of LC.

Results:  The results revealed that LC extract (25–100 µg/mL) concentration‐dependently decreased the PDGF‐induced cell proliferation as well as α‐smooth muscle actin expression in HSC. The inhibitory activity of LC on HSC was associated with: (i) inhibition of BrdU incorporation; (ii) induction of apoptosis with the activation of caspase‐3, up‐regulation of cell cycle inhibitory proteins p21 and p27, and down‐regulation of cell cycle stimulatory proteins cyclins D1 and D2; and (iii) increased phosphorylation of mitogen‐activated protein kinases (JNK). LC at the studied concentrations showed no direct cytotoxicity on primary hepatocytes.

Conclusion:  The results suggest that LC significantly inhibited PDGF‐activated HSC proliferation, possibly through apoptotic mechanisms and the potential of LC as an antifibrotic agent warrants further investigation.
Concurrent alterations of O‐GlcNAcylation and phosphorylation of tau in mouse brains during fasting
Authors: Li Xu , Lu Fen , Wang Jian‐Zhi , Gong Cheng‐Xin
Published: April 2006

Impaired brain glucose uptake/metabolism precedes the symptoms of Alzheimer disease (AD) and is likely to play a role in the development of the disease, but the mechanism by which it contributes to AD is not understood. Because glucose uptake/metabolism regulates protein O‐GlcNAcylation, and the latter modulates phosphorylation of tau inversely, we investigated, in fasting Kunming mice, whether impaired brain glucose uptake/metabolism causes abnormal hyperphosphorylation of tau and, consequently, facilitates the neurofibrillary degeneration of AD via down‐regulation of tau O‐GlcNAcylation. We found that fasting caused decreased tau O‐GlcNAcylation and concurrent hyperphosphorylation of tau at most of the phosphorylation sites studied. The hippocampus was found more vulnerable to the tau alterations than the cerebral cortex, which is consistent with the fact that it is the hippocampus that is first affected in AD. Furthermore, hyperphosphorylation of tau induced by fasting was reversible in the brain after re‐feeding. These findings provide a novel mechanism explaining how impaired brain glucose uptake/metabolism contributes to AD and suggest that it may be feasible to treat AD by reversing the abnormal hyperphosphorylation of tau at early stages of the disease.
Pro‐opiomelanocortin colocalizes with corticotropin‐ releasing factor in axon terminals of the noradrenergic nucleus locus coeruleus
Authors: Reyes Beverly A. S. , Glaser Julia D. , Magtoto Ronaldo , Van Bockstaele Elisabeth J.
Published: April 2006

We previously demonstrated that the opioid peptide enkephalin and corticotropin‐releasing factor (CRF) are occasionally colocalized in individual axon terminals but more frequently converge on common dendrites in the locus coeruleus (LC). To further examine potential opioid cotransmitters in CRF afferents we investigated the distribution of pro‐opiomelanocortin (POMC), the precursor that yields the potent bioactive peptide β‐endorphin, with respect to CRF immunoreactivity using immunofluorescence and immunoelectron microscopic analyses of the LC. Coronal sections were collected through the dorsal pontine tegmentum of rat brain and processed for immunocytochemical detection of POMC and CRF or tyrosine hydroxylase (TH). POMC‐immunoreactive processes exhibited a distinct distribution within the LC as compared to the enkephalin family of opioid peptides. Specifically, POMC fibers were enriched in the ventromedial aspect of the LC with fewer fibers present dorsolaterally. Immunofluorescence microscopy showed frequent coexistence of POMC and CRF in varicose processes that overlapped TH‐containing somatodendritic processes in the LC. Ultrastructural analysis showed POMC immunoreactivity in unmyelinated axons and axon terminals. Axon terminals containing POMC were filled with numerous large dense‐core vesicles. In sections processed for POMC and TH, ∼ 29% of POMC‐containing axon terminals (n = 405) targeted dendrites that exhibited immunogold–silver labeling for TH. In contrast, sections processed for POMC and CRF showed that 27% of POMC‐labeled axon terminals (n = 657) also exhibited CRF immunoreactivity. Taken together, these data indicate that a subset of CRF afferents targeting the LC contain POMC and may be positioned to dually impact LC activity.
Delayed ischemic electrocortical suppression during rapid repeated cerebral ischemia and kainate‐induced seizures in rat
Authors: Ilie Andrei , Spulber Stefan , Avramescu Sinziana , Nita Dragos Alexandru , Zagrean Ana‐Maria , Zagrean Leon , Moldovan Mihai
Published: April 2006

Global cerebral ischemia induces, within seconds, suppression of spontaneous electrocortical activity, partly due to alterations in synaptic transmission. In vitro studies have found that repeated brief hypoxic episodes prolong the persistence of synaptic transmission due to weakened adenosine release. The aim of this study was to investigate in vivo whether the time to ischemic electrocortical suppression (TES) could be altered during energy stress conditions such as rapid repeated global cerebral ischemia and kainate‐induced seizures. Experiments were carried out in adult rats under chloral hydrate anaesthesia. Repeated episodes of 1 min of ischemia were induced by transiently clamping the carotid arteries in a ‘four‐vessel occlusion’ model. We devised an automatic method of TES estimation based on the decay of the root mean square of two‐channel electrocorticographic recordings. To distinguish the alterations in spontaneous electrocortical activity we compared TES with the ischemic suppression of visual evoked potentials (VEP). During the first ischemic episode, TES was ∼ 15 s and remained unchanged when five ischemic episodes were separated by 10‐min reperfusion intervals. When ischemia was repeated after 2 min of reperfusion TES progressively increased, reaching a plateau value of ∼ 24 s. A similar plateau was reached during kainate‐induced seizures. The TES plateau occurred prior to ischemic suppression of VEP. Our data suggest that, under conditions of acute metabolic stress in vivo, the ischemic suppression of spontaneous electrocortical activity may be delayed up to a plateau value. These findings are consistent with the hypothesis of a depletable adenosine pool; however, the restoration of synaptic transmission may be faster in vivo than in vitro.