bluuebunny/crossref_metadata_2025 huggingface dataset:
First Report of Aspergillus niger as a Plant Pathogen on Zingiber officinale from India
Authors: Pawar N. V. , Patil V. B. , Kamble S. S. , Dixit G. B.
Published: August 2008

Zingiber officinale Rosc., belonging to the Zingiberaceae family, is medicinally important. It is commonly known as ginger and has been extensively cultivated for many centuries for use as a spice and traditional medicine in India. During a survey in September of 2007, leaves of Z. officinale showed a necrotic leaf spot disease on a plant growing in Kolhapur, India. Symptoms of the disease appeared as small (10 to 15 mm), rectangular to irregular, yellow spots on the leaves that covered a major area of the leaf when severe. The infection caused defoliation. Symptomatic leaves were collected and isolations from infected leaves were made on Czapek's Dox agar supplemented with streptomycin sulfate (30 mg/l). Plates were incubated at 28 to 30°C for 6 to 7 days. The fungal colonies were colorless to pale on the reverse side and covered with a dense layer of dark brown-to-black conidial heads. Conidia were globose to subglobose (3.5 to 5.0 μm in diameter), dark brown to black, and rough walled. 16S rRNA of isolates was amplified and sequenced (EMBL Accession No. AM941157) and compared with sequences of known Aspergillus species obtained from GenBank. The closest matches (99% identity) were with A. niger. On the basis of morphological and molecular approaches, the pathogen was confirmed as A. niger. Of five isolates, AN-5 was used for pathogenicity study. Koch's postulates were satisfied after reisolating the fungus from leaves inoculated with a conidial suspension that showed symptoms (6 days after inoculation) similar to the lesions observed on the leaves collected from Z. officinale. The tests were repeated three times in the greenhouse. Healthy, potted Z. officinale plants were grown in isolation after artificial inoculation with a suspension of spores (10,000 conidia per ml), which was prepared in 100 ml of sterile distilled water and then sprayed on the abaxial and adaxial surface of leaves. Noninoculated plants served as controls. These results confirmed the pathogenicity of A. niger on Z. officinale. To our knowledge, this is the first report of A. niger causing a disease on ginger (1).

Reference: (1) G. N. Dake. J. Spices Aromatic Crops. 4:40, 1995.
First Report of Powdery Mildew Caused by Oidium neolycopersici on Tomato in China
Authors: Li C. W. , Pei D. L. , Wang W. J. , Ma Y. S. , Wang L. , Wang F. , Liu J. L. , Zhu W. M.
Published: August 2008

Tomato powdery mildew can cause remarkable reduction in fruit size and quality (4). In March of 2008, powdery mildew appeared as circular, white colonies on leaves, petioles, and stems of tomato plants grown in greenhouses in Shangqiu, Henan Province, China. The pathogenic fungus had unbranched conidiophores with an average length of 58.4 μm and width of 5.1 μm. Conidia were hyaline, elliptical, and were borne singly. Average length and width of conidia were 30.6 and 15.1 μm, respectively. Germ tubes were straight and formed at the ends or very close to the ends of conidia. Chasmothecium was not found in the collected samples. Different tomato cultivars and species, including Lycopersicon esculentum Mill (cvs. Moneymaker, Micro-Tom, Zaofen, Fenguo, and Zhongza series), L. peruvianum cv. LA2172, and L. hirsutum cv. G1.1560, were inoculated with a conidial suspension with a concentration of 5 × 104 conidia/ml. Plants developed powdery mildew symptoms as early as 4 days after inoculation. Susceptible symptoms developed on all L. esculentum cultivars, while L. peruvianum LA2172 and L. hirsutum G1.1560 displayed complete resistance, which is similar to the results of Bai et al 2004 (1) and Lindhout and Pet 1990 (3). Morphological characteristics of the pathogen on susceptible genotypes were similar to those from naturally infected plants. On the basis of the characteristics of the asexual stage, the pathogen was identified as an isolate of Oidium neolycopersici L. Kiss, which was confirmed by internal transcribed spacer (ITS) sequence analysis. PCR amplification and sequencing of the ITS region were performed with primers ITS1 and ITS4. The nucleotide sequence was assigned GenBank Accession No. EU486992, which had a 100% homology with 10 ITS sequences of O. neolycopersici in GenBank (Accession Nos. EU047559 to 047568) (2). In Asia, the spread of this pathogen has been recently reported in Japan (2). To our knowledge, this is the first report of tomato powdery mildew in China. Voucher specimens are available at the Specimen Center in the Department of Life Science, Shangqiu Normal University.

References: (1) Y. Bai et al. Mol. Plant-Microbe. Interact. 18:354, 2005. (2) T. Jankovics et al. Phytopathology 98:529, 2008. (3) P. Lindhout and G. Pet. Tomato Gen. Coop. Rep. 40:19, 1990. (4) J. M. Whipps et al. Plant Pathol. 47:36, 1998.
First Report of Fulvia fulva, Causal Agent of Tomato Leaf Mold, in Greenhouses in Southeastern Spain
Authors: de Cara M. , Heras F. , Santos M. , Marquina J. C. Tello
Published: August 2008

Tomato (Solanum lycopersicum L.) is produced in more than 9,000 ha of greenhouses in Almería (southeastern Spain). During 2006 and 2007, a new disease was observed on almost all plants in 37 greenhouses. Yellow spots on upper and lower leaf surfaces were accompanied by gray-to-dark brown mycelia, conidiophores, and conidia on lower leaf surfaces. Affected leaves became necrotic and withered. Six isolates grown on malt extract agar (MEA) were identified as Fulvia fulva (1). The one- to three-celled conidia ranged from 21.8 × 7.8 μm to 21.5 × 6.5 μm. On MEA, potato dextrose agar, and V8 juice agar, the pathogen grew slowly; colonies were only 1 cm in diameter after 30 days. Colony color was initially intense yellow but became dark brown with age. In a growth chamber (12,000 lux for 16 h per day, 23 to 28°C, and 60 to 95% relative humidity), six pots containing five tomato plants (cv. SanPedro) at the four-true-leaf stage were inoculated with a conidial suspension (103 CFU/ml) of F. fulva. Control plants were sprayed with water. The trial was repeated once. Immediately after inoculation, plants were sealed in plastic bags for 8 days. Symptoms of the disease and signs of the pathogen were observed on all inoculated plants 18 days after inoculation. To our knowledge, this is the first report of leaf mold of tomato in Almería and it is becoming common in the greenhouse industry in this region.

Reference: (1) P. Holliday and J. L. Mulder. No. 487 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1976.
Genetic Relationships Among Populations of Gibberella zeae from Barley, Wheat, Potato, and Sugar Beet in the Upper Midwest of the United States
Authors: Burlakoti R. R. , Ali S. , Secor G. A. , Neate S. M. , McMullen M. P. , Adhikari T. B.
Published: August 2008

Gibberella zeae, a causal agent of Fusarium head blight (FHB) in wheat and barley, is one of the most economically harmful pathogens of cereals in the United States. In recent years, the known host range of G. zeae has also expanded to noncereal crops. However, there is a lack of information on the population genetic structure of G. zeae associated with noncereal crops and across wheat cultivars. To test the hypothesis that G. zeae populations sampled from barley, wheat, potato, and sugar beet in the Upper Midwest of the United States are not mixtures of species or G. zeae clades, we analyzed sequence data of G. zeae, and confirmed that all populations studied were present in the same clade of G. zeae. Ten variable number tandem repeat (VNTR) markers were used to determine the genetic structure of G. zeae from the four crop populations. To examine the effect of wheat cultivars on the pathogen populations, 227 strains were sampled from 10 subpopulations according to wheat cultivar types. The VNTR markers also were used to analyze the genetic structure of these subpopulations. In all populations, gene (H = 0.453 to 0.612) and genotype diversity (GD = >0.984) were high. There was little or no indication of linkage disequilibrium (LD) in all G. zeae populations and subpopulations. In addition, high gene flow (Nm) values were observed between cereal and noncereal populations (Nm = 10.69) and between FHB resistant and susceptible wheat cultivar subpopulations (Nm = 16.072), suggesting low population differentiation of G. zeae in this region. Analysis of molecular variance also revealed high genetic variation (>80%) among individuals within populations and subpopulations. However, low genetic variation (<5%) was observed between cereal and noncereal populations and between resistant and susceptible wheat subpopulations. Overall, these results suggest that the populations or subpopulations are likely a single large population of G. zeae affecting crops in the upper Midwest of the United States.
Seed-Colonizing Microbes from Municipal Biosolids Compost Suppress Pythium ultimum Damping-Off on Different Plant Species
Authors: Chen M.-H. , Nelson E. B.
Published: August 2008

Composts are known for their suppressive properties toward many different seed- and root-infecting pathogens and diseases. Although disease and pathogen suppression induced by composts is believed to be mediated by microbial activities, the nature of the microbial species and processes responsible for suppressiveness remain unknown. We demonstrated previously that seed-colonizing microbial consortia from leaf compost could explain the observed levels of Pythium ultimum-induced damping-off suppression on cotton. The aim of the present work was to determine whether seed-colonizing microbial consortia could explain Pythium damping-off suppression in municipal biosolids compost on three different plant species. Significant levels of disease suppression were observed on cucumber, wheat, and pea at water potentials of –2 kPa. The suppression of damping-off on cucumber and wheat could be eliminated by autoclaving the compost prior to sowing. High levels of suppressiveness were expressed both on cucumber and on wheat seed surfaces within 8 h of sowing. However, the expression of damping-off suppression on the surface of pea seeds was inconsistent and highly variable. Our results demonstrate that compost-induced suppression of P. ultimum damping-off of cucumber and wheat can be explained by the microbial consortia colonizing seeds within 8 h of sowing. These results further suggest that disease suppression in composts is related to microbial species that interact with the pathogen in its infection court and not in the bulk compost.
Phylogenetic Analysis of Verticillium dahliae Vegetative Compatibility Groups
Authors: Collado-Romero M. , Mercado-Blanco J. , Olivares-García C. , Jiménez-Díaz R. M.
Published: August 2008

The evolutionary relationships among Verticillium dahliae vegetative compatibility (VCG) subgroups VCG1A, VCG1B, VCG2A, VCG2B, VCG4A, VCG4B, and VCG6 were investigated by parsimony analysis of amplified fragment length polymorphism (AFLP) fingerprints and sequences of six DNA regions (actin, β-tubulin, calmodulin, and histone 3 genes, the ITS 1 and 2 regions of the rDNA, and a V. dahliae-specific sequence), using 101 isolates of diverse host and geographic origin. Polymorphisms in gene sequences among isolates of different VCGs were very low and individual gene genealogies provided very little resolution at the VCG level. The combined analysis of all DNA regions differentiated all VCG subgroups except for isolates in VCG1A and VCG1B. VCG clonal lineages in V. dahliae and evolutionary relationships among them were resolved independently by analyses of AFLP fingerprints, multiple gene genealogies, and the combined data set of AFLP fingerprinting and multiple gene genealogies. Two main lineages (I and II) were identified with lineage II comprising two closely related subgroups of VCGs. Lineage I included VCG1A, VCG1B, and VCG2B334; and lineage II included, VCG2A and VCG4B (subclade 1); and VCG2B824, VCG4A, and VCG6 (subclade 2). VCG subgroups were monophyletic except for VCG2B that appeared polyphyletic. Limiting the parsimony analysis either to AFLP fingerprints or DNA sequences would have obscured intra-VCG differentiation. Therefore, the dual approach represented by the independent and combined analyses of AFLP fingerprints and DNA sequences was a highly valuable method for the identification of phylogenetic relationships at the intraspecific level in V. dahliae.
Amino Acid Polymorphisms in Strictly Conserved Domains of a P-Type ATPase HMA5 Are Involved in the Mechanism of Copper Tolerance Variation in Arabidopsis
Authors: Kobayashi Yuriko , Kuroda Keishi , Kimura Keisuke , Southron-Francis Jennafer L. , Furuzawa Aya , Kimura Kazuhiko , Iuchi Satoshi , Kobayashi Masatomo , Taylor Gregory J. , Koyama Hiroyuki
Published: August 2008

Copper (Cu) is an essential element in plant nutrition, but it inhibits the growth of roots at low concentrations. Accessions of Arabidopsis (Arabidopsis thaliana) vary in their tolerance to Cu. To understand the molecular mechanism of Cu tolerance in Arabidopsis, we performed quantitative trait locus (QTL) analysis and accession studies. One major QTL on chromosome 1 (QTL1) explained 52% of the phenotypic variation in Cu tolerance in roots in a Landsberg erecta/Cape Verde Islands (Ler/Cvi) recombinant inbred population. This QTL regulates Cu translocation capacity and involves a Cu-transporting P1B-1-type ATPase, HMA5. The Cvi allele carries two amino acid substitutions in comparison with the Ler allele and is less functional than the Ler allele in Cu tolerance when judged by complementation assays using a T-DNA insertion mutant. Complementation assays of the ccc2 mutant of yeast using chimeric HMA5 proteins revealed that N923T of the Cvi allele, which was identified in the tightly conserved domain N(x)6YN(x)4P (where the former asparagine was substituted by threonine), is a cause of dysfunction of the Cvi HMA5 allele. Another dysfunctional HMA5 allele was identified in Chisdra-2, which showed Cu sensitivity and low capacity of Cu translocation from roots to shoots. A unique amino acid substitution of Chisdra-2 was identified in another strictly conserved domain, CPC(x)6P, where the latter proline was replaced with leucine. These results indicate that a portion of the variation in Cu tolerance of Arabidopsis is regulated by the functional integrity of the Cu-translocating ATPase, HMA5, and in particular the amino acid sequence in several strictly conserved motifs.
Combined Genetic and Modeling Approaches Reveal That Epidermal Cell Area and Number in Leaves Are Controlled by Leaf and Plant Developmental Processes in Arabidopsis
Authors: Tisné Sébastien , Reymond Matthieu , Vile Denis , Fabre Juliette , Dauzat Myriam , Koornneef Maarten , Granier Christine
Published: August 2008

Both leaf production and leaf expansion are tightly linked to cell expansion and cell division, but the functional relationships between all these variables are not clearly established. To get insight into these relationships, a quantitative genetic analysis was performed in 118 recombinant inbred lines derived from a cross between the Landsberg erecta and Antwerp accessions and was combined with a structural equation modeling approach. Main effects and epistatic interactions at the quantitative trait locus (QTL) level were detected for rosette area, rosette leaf number, leaf 6 area, epidermal cell area and number. A QTL at ERECTA marker (ER) controlled cell expansion and cell division, in interaction with two other QTLs at SNP295 and SNP21 markers. Moreover, both the screening for marker association involved in the variation of the relationships between leaf growth variables and the test of alternative functional models by structural equation modeling revealed that the allelic value at ER controlled epidermal cell area and epidermal cell number in a leaf. These effects are driven both by a whole plant mechanism associated with leaf production and by a single leaf mechanism associated with leaf expansion. The complex effects of the QTL at ER were validated in selected heterogeneous inbred families. The ERECTA gene, which is mutated in the Landsberg erecta parental line, was found to be a putative candidate responsible for these mapped effects by phenotyping mutants of this gene at the cellular level. Together, these results give insight into the complex determination of leaf epidermal cell number and area.
Lysophosphatidic acid stimulates the proliferation and motility of malignant pleural mesothelioma cells through lysophosphatidic acid receptors, LPA1 and LPA2
Authors: Yamada Tadaaki , Yano Seiji , Ogino Hirokazu , Ikuta Kenji , Kakiuchi Soji , Hanibuchi Masaki , Kanematsu Takanori , Taniguchi Tetsuo , Sekido Yoshitaka , Sone Saburo
Published: August 2008

Lysophosphatidic acid (LPA) is one of the simplest natural phospholipids. This phospholipid is recognized as an extracellular potent lipid mediator with diverse effects on various cells. Although LPA is shown to stimulate proliferation and motility via LPA receptors, LPA1 and LPA2, in several cancer cell lines, the role of LPA and LPA receptors for malignant pleural mesothelioma (MPM) has been unknown. MPM is an aggressive malignancy with a poor prognosis and the incidence is increasing and is expected to increase further for another 10–20 years worldwide. Therefore, the development of novel effective therapies is needed urgently. In this study, we investigated the effect of LPA on the proliferation and motility of MPM cells. We found that all 12 cell lines and four clinical samples of MPM expressed LPA1, and some of them expressed LPA2, LPA3, LPA4 and LPA5. LPA stimulated the proliferation and motility of MPM cells in a dose‐dependent manner. Moreover, LPA‐induced proliferation was inhibited by Ki16425, an inhibitor of LPA1, and small interfering RNA against LPA1, but not LPA2. Interestingly, LPA‐induced motility was inhibited by small interfering RNA against LPA2, but not LPA1, unlike a number of previous reports. These results indicate that LPA is a critical factor on proliferation though LPA1, and on motility though LPA2 in MPM cells. Therefore, LPA and LPA receptors, LPA2 as well as LPA1, represent potential therapeutic targets for patients with MPM. (Cancer Sci 2008; 99: 1603–1610)
Enhancement of antiproliferative activity of interferons by RNA interference‐mediated silencing of SOCS gene expression in tumor cells
Authors: Takahashi Yuki , Kaneda Haruka , Takasuka Nana , Hattori Kayoko , Nishikawa Makiya , Takakura Yoshinobu
Published: August 2008

The suppressor of cytokine signaling (SOCS) proteins, negative regulators of interferon (IFN)‐induced signaling pathways, is involved in IFN resistance of tumor cells. To improve the growth inhibitory effect of IFN‐β and IFN‐γ on a murine melanoma cell line, B16‐BL6, and a murine colon carcinoma cell line, Colon26 cells, SOCS‐1 and SOCS‐3 gene expression in tumor cells was downregulated by transfection of plasmid DNA expressing short hairpin RNA targeting one of these genes (pshSOCS‐1 and pshSOCS‐3, respectively). Transfection of pshSOCS‐1 significantly increased the antiproliferative effect of IFN‐γ on B16‐BL6 cells. However, any other combinations of plasmids and IFN had little effect on the growth of B16‐BL6 cells. In addition, transfection of pshSOCS‐1 and pshSOCS‐3 produced little improvement in the effect of IFN on Colon26 cells. To understand the mechanism underlining these findings, the level of SOCS gene expression was measured by real time polymerase chain reaction. Addition of IFN‐γ greatly increased the SOCS‐1 mRNA expression in B16‐BL6 cells. Taking into account the synergistic effect of pshSOCS‐1 and IFN‐γ on the growth of B16‐BL6 cells, these findings suggest that IFN‐γ‐induced high SOCS‐1 gene expression in B16‐BL6 cells significantly interferes with the antiproliferative effect of IFN‐γ. These results indicate that silencing SOCS gene expression can be an effective strategy to enhance the antitumor effect of IFN under conditions in which the SOCS gene expression is upregulated by IFN. (Cancer Sci 2008; 99: 1650–1655)
Estimation of the relationship between caspase‐3 expression and clinical outcome of Burkitt's and Burkitt‐like lymphoma
Authors: Nomura Yuko , Yoshida Shiro , Karube Kennosuke , Takeshita Morishige , Hirose Shinichi , Nakamura Shigeo , Yoshino Tadashi , Kikuchi Masahiro , Ohshima Koichi
Published: August 2008

Burkitt's lymphoma and atypical Burkitt/Burkitt‐like lymphoma (BL/BLL) are considered highly aggressive B‐cell lymphomas with a rapid proliferative rate and high rate of apoptosis. The aim of the present study was to confirm whether apoptotic and cell proliferative factors affect BL/BLL clinical outcomes. We retrospectively analyzed the relationship between the clinical and immunophenotypic features of 43 BL/BLL patients by immunohistochemical staining for bcl‐2 and double staining for Ki‐67 plus caspase‐3. In double staining experiments, all patients were divided into high and low groups for the expression of caspase‐3, Ki‐67, and both Ki‐67 and caspase‐3, by using the medians of their percentages as limits. The 43 BL/BLL patients were divided into high caspase‐3 (n = 19) and low caspase‐3 (n = 24) groups. There was a significant difference in the overall survival between the high (77%) and low caspase‐3 (33%) groups; the survival rate of patients in the low caspase‐3 group who received aggressive short‐term chemotherapy (58%) was significantly better than that of patients who received cyclophosphamide, doxorubicin, vincristine and prednisolone (CHOP) therapy (17%). All patients positive for bcl‐2 were in the low caspase‐3 group (high caspase‐3 group, 0%; low caspase‐3 group, 42%). The overall survival tended to be better in the high caspase‐3 and bcl‐2‐negative group (76%) than in the low caspase‐3 and bcl‐2‐negative (50%) group. In addition, the low caspase‐3 and bcl‐2‐positive group tended to show the worst prognosis (16%). We suggest that caspase‐3 may function as an indicator of the prognosis of BL/BLL. Furthermore, intensive short‐term chemotherapeutic regimens may improve the prognosis of the patients in the low caspase‐3 group. (Cancer Sci 2008; 99: 1564–1569)
Molecular links between tumor angiogenesis and inflammation: inflammatory stimuli of macrophages and cancer cells as targets for therapeutic strategy
Authors: Ono Mayumi
Published: August 2008

Both inflammation and angiogenesis are exacerbated by increased production of chemokines/cytokines, growth factors, proteolytic enzymes, proteoglycans, lipid mediators and prostaglandins. It has been reported that approximately 15–20% of all malignancies are initiated or exacerbated by inflammation. Initiation and progression of cancer are also closely linked to angiogenesis. Infiltration of macrophages is a dramatic and common feature of inflammation, angiogenesis and cancer, and has been recently highlighted in an attempt to develop novel strategies for treating cancer. The recruitment and infiltration of macrophages in the tumor microenvironment activates them to support the malignant progression of cancer cells, and these macrophages are called tumor‐associated macrophages. In a model of experimental angiogenesis using mouse corneas, macrophages infiltrated tissue in response to inflammatory cytokines and produced chemokines and angiogenesis‐promoting factors, such as vascular endothelial growth factor‐A, interleukin‐8, matrix metalloproteinases, prostanoids and reactive oxygen species. Moreover, in a cancer xenograft model, inflammatory stimuli by a representative inflammatory cytokine, interleukin‐1β, enhanced tumor growth and angiogenesis with infiltration and activation of macrophages. Co‐culture of cancer cells with macrophages synergistically stimulated production of various angiogenesis‐related factors when stimulated by the inflammatory cytokine. This inflammatory angiogenesis in both mouse cornea and a tumor model was mediated, in part, by activation of nuclear factor κB and activator protein 1 (Jun/Fos). Administration of either nuclear factor κB‐targeting drugs or cyclooxygenase 2 inhibitors or depletion of macrophages could block both inflammatory angiogenesis and tumor angiogenesis. Thus, both inflammatory and angiogenic responses in tumor stroma could be targets for development of anticancer therapeutic drugs. (Cancer Sci 2008; 99: 1501–1506)
Humoral immune responses against tumor‐associated antigen OVA66 originally defined by serological analysis of recombinant cDNA expression libraries and its potentiality in cellular immunity
Authors: Jin Shu , Wang Ying , Zhang Yong , Zhang Hui‐Zhen , Wang Shu‐Jun , Tang Jun‐Qiao , Chen Hui‐Juan , Ge Hai‐Liang
Published: August 2008

Immunotherapy for cancer relies on the identification of tumor antigens and efficacy of antitumor immune responses. Serological analysis of recombinant cDNA libraries (SEREX), which is based on the spontaneous humoral responses against potential tumor antigens, has provided a novel strategy for searching novel tumor‐associated candidates. Through SEREX analysis, we have identified 24 distinct gene clones by immunoscreening of a cDNA library derived from an ovarian cancer patient. Among these genes, a novel gene, OVA66, was found to be expressed significantly higher in carcinoma samples from cancer patients than in normal controls. Comparing humoral responses to OVA66 between tumor patients and healthy donors, it has been shown that the IgG level against OVA66 was significantly elevated in the serum of cancer patients from different histological types of cancer. To determine whether SEREX‐defined OVA66 can trigger promising cytotoxic T lymphocyte (CTL) responses, human leukocyte antigen (HLA)‐A*0201‐restricted T‐cell epitopes were predicted through a computational algorithm. Of four predicted peptides, p306–314 (L235) possesses the ability to induce efficient peripheral blood lymphocyte (PBL)‐derived CTL responses capable of specifically recognizing peptide‐pulsed T2 cells and lysing carcinoma cell lines expressing both HLA‐A2 and OVA66 as determined by cytotoxicity and enzyme‐linked immunospot assay (ELISPOT). Taken together, our results demonstrate that the SEREX‐defined tumor‐associated antigen OVA66 can elicit humoral immunity and may also serve as a potential candidate for T‐cell‐based immunotherapy for cancer. (Cancer Sci 2008; 99: 1670–1678)
Expression and mutation analysis of epidermal growth factor receptor in head and neck squamous cell carcinoma
Authors: Ali Mahmoud A.L. Sheikh , Gunduz Mehmet , Nagatsuka Hitoshi , Gunduz Esra , Cengiz Beyhan , Fukushima Kunihiro , Beder Levent Bekir , Demircan Kadir , Fujii Masae , Yamanaka Noboru , Shimizu Kenji , Grenman Reidar , Nagai Noriyuki
Published: August 2008

The epidermal growth factor receptor (EGFR)–RAS–RAF–mitogen‐activated protein kinase signaling cascade is an important pathway in cancer development and recent reports show that EGFR and its downstream signaling molecules are mutated in a number of cancers. We have analyzed 91 Japanese head and neck squamous cell carcinomas (HNSCC) and 12 HNSCC cell lines for mutations in EGFR, ErbB2, and K‐ras. Exons encoding the hot‐spot regions in the tyrosine kinase domain of both EGFR (exons 18, 19, and 21) and ErbB2 (exons 18–23), as well as exons 1 and 2 of K‐ras were amplified by polymerase chain reaction and sequenced directly. EGFR expression was also analyzed in 65 HNSCC patients using immunohistochemistry. Only one silent mutation, C836T, was found in exon 21 of EGFR in the UT‐SCC‐16A cell line and its corresponding metastasic cell line UT‐SCC‐16B. No other mutation was found in EGFR, ErbB2, or K‐ras. All tumors showed EGFR expression. In 21 (32%) tumors, EGFR was expressed weakly (+1). In 27 (42%) tumors it was expressed (+2) moderately, and in 17 (26%) tumors high expression (+3) was detected. Overexpression (+2, +3) was found in 44 tumors (68%). A worse tumor differentiation and a positive nodal stage were significantly associated with EGFR overexpression (P = 0.02, P = 0.032, respectively). Similar to patients from western ethnicity, mutations are absent or rare in Japanese HNSCC. Protein overexpression rather than mutation might be responsible for activation of the EGFR pathway in HNSCC. (Cancer Sci 2008; 99: 1589–1594)
Caffeine yields aneuploidy through asymmetrical cell division caused by misalignment of chromosomes
Authors: Katsuki Yoko , Nakada Shinichiro , Yokoyama Tetsuji , Imoto Issei , Inazawa Johji , Nagasawa Masayuki , Mizutani Shuki
Published: August 2008

Aneuploidy has been implicated as an important step leading to various neoplasias. Although genetic factors that block aneuploidy have been the subject of intense interest, the impact of pharmacological and environmental substances on the development of aneuploidy has not been studied. Here, we show that caffeine induces aneuploidy through asymmetrical cell division. Mitotic exits of HeLa, U2OS, and primary fibroblast cells were significantly delayed by 10 mmol/L caffeine. Most caffeine‐treated mitotic cells showed misalignment of chromosomes at the metaphase plates, and were arrested at prometaphase. Mitoticarrest deficient 2 (MAD2) depletion rescued the caffeine‐induced delay of mitotic exit, indicating that caffeine‐induced prolongation of mitosis was caused by activation of a MAD2‐dependent spindle checkpoint. Enumeration of centromeres by fluorescence in situ hybridization revealed that cell division in the presence of caffeine was not symmetrical and resulted in aneuploid cell production. Most of these cells survived and underwent DNA synthesis. Our findings reveal a novel pharmacological effect of a high concentration of caffeine on genomic stability in dividing cells. (Cancer Sci 2008; 99: 1539–1545)
Head and neck cancer in the betel quid chewing area: recent advances in molecular carcinogenesis
Authors: Chen Yin‐Ju , Chang Joseph Tung‐Chieh , Liao Chun‐Ta , Wang Hung‐Ming , Yen Tzu‐Chen , Chiu Ching‐Chi , Lu Ya‐Ching , Li Hsiao‐Fang , Cheng Ann‐Joy
Published: August 2008

Head and neck cancer (HNC) is one of the 10 most frequent cancers worldwide, with an estimated over 500 000 new cases being diagnosed annually. The overall 5‐year survival rate in patients with HNC is one of the lowest among common malignant neoplasms and has not significantly changed during the last two decades. Oral cavity squamous cell carcinoma (OSCC) shares part of HNC and has been reported to be increasing in the betel quid chewing area in recent years. During 2006, OSCC has become the sixth most common type of cancer in Taiwan, and it is also the fourth most common type of cancer among men. It follows that this type of cancer wreaks a high social and personal cost. Environmental carcinogens such as betel quid chewing, tobacco smoking and alcohol drinking have been identified as major risk factors for head and neck cancer. There is growing interest in understanding the relationship between genetic susceptibility and the prevalent environmental carcinogens for HNC prevention. Within this review, we discuss the molecular and cellular aspects of HNC carcinogenesis in Taiwan, an endemic betel quid chewing area. Knowledge of molecular carcinogenesis of HNC may provide critical clues for diagnosis, prognosis, individualization of therapy and molecular therapeutics. (Cancer Sci 2008; 99: 1507–1514)
Roles of the histone acetyltransferase monocytic leukemia zinc finger protein in normal and malignant hematopoiesis
Authors: Katsumoto Takuo , Yoshida Naomi , Kitabayashi Issay
Published: August 2008

Histone‐modified enzymes are involved in various cell functions, including proliferation, differentiation, cell death and carcinogenesis. The protein MOZ (monocytic leukemia zinc finger protein) is a Myst (MOZ, Ybf2 (Sas3), Sas2, Tip60)‐type histone acetyltranseferase (HAT) that generates fusion genes, such as MOZ–TIF2, MOZ–CBP and MOZ–p300, in acute myeloid leukemia (AML) by chromosomal translocation. MOZ associates with AML1 (RUNX1), PU.1, and p53, and cooperatively activates target gene transcription. Gene targeting in mice has revealed that MOZ is essential for the generation and maintenance of hematopoietic stem cells (HSC) and for the appropriate development of myeloid, erythroid and B‐lineage cell progenitors. In AML, MOZ fusion genes lead to repressed differentiation, hyper‐proliferation, and self‐renewal of myeloid progenitors through deregulation of MOZ‐regulated target gene expression. It is therefore necessary to analyze the roles of MOZ and MOZ fusion genes in normal and malignant hematopoiesis to elucidate the mechanisms underlying and develop therapies for MOZ‐related AML. (Cancer Sci 2008; 99: 1523–1527)
Erythropoietin‐producing hepatocyte B6 variant‐derived peptides with the ability to induce glioma‐reactive cytotoxic T lymphocytes in human leukocyte antigen‐A2+ glioma patients
Authors: Jin Mingyue , Komohara Yoshihiro , Shichijo Shigeki , Yamanaka Ryuya , Nikawa Junichi , Itoh Kyogo , Yamada Akira
Published: August 2008

We recently cloned a variant form of erythropoietin‐producing hepatocyte (Eph)B6, a member of the Eph receptor tyrosine kinase family. In the present study, we examined the expression of the EphB6 variant (EphB6v) in a panel of brain tumor cell lines and glioblastoma tissues and we found that EphB6v was preferentially expressed in malignant brain tumors, such as glioblastomas and anaplastic astrocytomas. The EphB6v has a unique 54 amino acid sequence at the C‐terminal that is not found in normal EphB6. Therefore, we attempted to identify antigenic peptides unique to EphB6v for immunotherapy. The two EphB6v‐derived peptides exhibited the ability to bind to human leukocyte antigen (HLA)‐A0201 molecules, and each of them was able to induce cytotoxic T lymphocytes in vitro in the peripheral blood mononuclear cells of HLA‐A2+ glioma patients. The cytotoxicity was mediated by peptide‐specific CD8+ T cells in an HLA‐A2‐restricted manner. The expression of EphB6v was also observed in different types of cancer (e.g. lung, colon, stomach, liver and pancreatic) cells. Taken together, the two peptides derived from EphB6v might be appropriate targets for peptide‐based specific immunotherapy for HLA‐A2+ patients with various cancers. (Cancer Sci 2008; 99: 1656–1662)
Up‐regulation of matrix metalloproteinase‐3 in the dorsal root ganglion of rats with paclitaxel‐induced neuropathy
Authors: Nishida Kentaro , Kuchiiwa Satoshi , Oiso Shigeru , Futagawa Toshitaka , Masuda Shogo , Takeda Yasuo , Yamada Katsushi
Published: August 2008

Paclitaxel‐induced painful peripheral neuropathy is a major dose‐limiting factor. Recently, it has been reported that macrophages accumulated in the dorsal root ganglion of paclitaxel‐treated rats, and their activation is suggested to contribute to generation and development of the neuropathy. However, the mechanism for macrophage activation is still unknown. In this study, to explore candidate genes involved in the mechanism for macrophage activation in the dorsal root ganglion of paclitaxel‐treated rats, we developed model rats for paclitaxel‐induced neuropathic pain and performed a microarray assay to analyze the changes of gene expressions in the dorsal root ganglion. Among the genes with changed expression levels, we focused on matrix metalloproteinase‐3 (MMP‐3, stromelysin‐1) and CD163, a macrophage marker. By reverse transcription–polymerase chain reaction, the expression levels of MMP‐3 and CD163 were markedly up‐regulated in paclitaxel‐treated dorsal root ganglion. As a result of immunohistochemical study, large ganglion neurons, but neither Schwann cells nor macrophages, predominantly expressed MMP‐3. This MMP‐3 up‐regulation occurred prior to macrophage accumulation in the dorsal root ganglion. In addition, recombinant MMP‐3 led to the activation of RAW264 macrophages in vitro. Taken together, the up‐regulation of MMP‐3 and following macrophage activation caused in the dorsal root ganglion might be a significant event to trigger a series of reactions developing paclitaxel‐induced peripheral neuropathic pain. (Cancer Sci 2008; 99: 1618–1625)
Treating adolescent drug abuse: a randomized trial comparing multidimensional family therapy and cognitive behavior therapy
Authors: Liddle Howard A. , Dakof Gayle A. , Turner Ralph M. , Henderson Craig E. , Greenbaum Paul E.
Published: August 2008

Aim To examine the efficacy of two adolescent drug abuse treatments: individual cognitive behavioral therapy (CBT) and multidimensional family therapy (MDFT).

Design A 2 (treatment condition) x 4 (time) repeated‐measures intent‐to‐treat randomized design. Data were gathered at baseline, termination, 6 and 12 months post‐termination. Analyses used latent growth curve modeling.

Setting Community‐based drug abuse clinic in the northeastern United States.

Participants A total of 224 youth, primarily male (81%), African American (72%), from low‐income single‐parent homes (58%) with an average age of 15 years were recruited into the study. All youth were drug users, with 75% meeting DSM‐IV criteria for cannabis dependence and 13% meeting criteria for abuse.

Measurements Five outcomes were measured: (i) substance use problem severity; (ii) 30‐day frequency of cannabis use; (iii) 30‐day frequency of alcohol use; (iv) 30‐day frequency of other drug use; and (v) 30‐day abstinence.

Findings Both treatments produced significant decreases in cannabis consumption and slightly significant reductions in alcohol use, but there were no treatment differences in reducing frequency of cannabis and alcohol use. Significant treatment effects were found favoring MDFT on substance use problem severity, other drug use and minimal use (zero or one occasion of use) of all substances, and these effects continued to 12 months following treatment termination.

Conclusion Both interventions are promising treatments. Consistent with previous controlled trials, MDFT is distinguished by the sustainability of treatment effects.
Work‐place social capital and smoking cessation: the Finnish Public Sector Study
Authors: Kouvonen Anne , Oksanen Tuula , Vahtera Jussi , Väänänen Ari , De Vogli Roberto , Elovainio Marko , Pentti Jaana , Leka Stavroula , Cox Thomas , Kivimäki Mika
Published: August 2008

Aims To examine whether high social capital at work is associated with an increased likelihood of smoking cessation in baseline smokers.

Design Prospective cohort study.

Setting Finland.

Participants A total of 4853 employees who reported to be smokers in the baseline survey in 2000–2002 (response rate 68%) and responded to a follow‐up survey on smoking status in 2004–2005 (response rate 77%).

Measurements Work‐place social capital was assessed using a validated and psychometrically tested eight‐item measure. Control variables included sex, age, socio‐economic position, marital status, place of work, heavy drinking, physical activity, body mass index and physician‐diagnosed depression.

Findings In multi‐level logistic regression models adjusted for all the covariates, the odds for being a non‐smoker at follow‐up were 1.26 [95% confidence interval (CI) = 1.03–1.55] times higher for baseline smokers who reported high individual‐level social capital than for their counterparts with low social capital. In an analysis stratified by socio‐economic position, a significant association between individual‐level social capital and smoking cessation was observed in the high socio‐economic group [odds ratio (OR) (95% CI) = 1.63 (1.01–2.63)], but not in intermediate [(OR = 1.10 (0.83–1.47)] or low socio‐economic groups [(OR = 1.28 (0.86–1.91)]. Work unit‐level social capital was not associated with smoking cessation.

Conclusions If the observed associations are causal, these findings suggest that high perceived social capital at work may facilitate smoking cessation among smokers in higher‐status jobs.
A Scotland‐wide pilot programme of smoking cessation services for young people: process and outcome evaluation
Authors: Gnich Wendy , Sheehy Christine , Amos Amanda , Bitel Mark , Platt Stephen
Published: August 2008

Aim  To conduct an independent, external evaluation of a Scotland‐wide youth cessation pilot programme, focusing upon service uptake and effectiveness.

Intervention  National Health Service (NHS) Health Scotland and Action on Smoking and Health (ASH) Scotland funded a 3‐year (2002–2005) national pilot programme comprising eight projects which aimed to engage with and support young smokers (aged 12–25 years) to quit.

Design, participants and measurements  Process evaluation was undertaken via detailed case studies comprising qualitative interviews, observation and documentary analysis. Outcomes were assessed by following project participants (n = 470 at baseline) at 3 and 12 months and measuring changes in smoking behaviour, including carbon monoxide (CO)‐validated quit status.

Findings  Recruitment proved difficult. Considerable time and effort were needed to attract young smokers. Advertising and recruitment had to be tailored to project settings and educational activities proved essential to raise the profile of smoking as an issue. Thirty‐nine participants [8.6%, 95% confidence interval (CI) 5.0–11.2%] were CO‐validated quitters at 3 months and 11 of these (2.4%, 95% CI 1.90–3.8%) were also validated quitters at 12 months. Older participants were more likely to be abstinent at 3 months.

Conclusions  The overall quit rate was disappointing. As a result of low participant numbers, it was impossible to draw conclusions about the relative effectiveness of different project approaches. These findings give little support to the case for developing dedicated youth cessation services in Scotland. They also highlight the difficulties of undertaking ‘real‐world’ evaluations of pilot youth cessation projects. More action is needed to develop environments which enhance young smokers' motivation to quit and their ability to sustain quit attempts.
Insulin effect on adipose tissue (AT) adiponectin expression is regulated by the insulin resistance status of the patients
Authors: Hernandez‐Morante J. J. , Milagro F. I. , Lujan J. A. , Martinez J. A. , Zamora S. , Garaulet M.
Published: August 2008

Objective  The objective of the present study was to determine a possible depot‐specific effect of insulin‐stimulation on adiponectin gene expression in adipose tissue (AT) explants from subcutaneous and visceral AT. A secondary aim was to analyse the associations of adiponectin plasma levels, as well as control and insulin‐stimulated gene expression levels with different features of the metabolic syndrome.

Design  Visceral and subcutaneous AT biopsies were obtained from 20 subjects (10 men and 10 women) with morbid obesity. Metabolic syndrome and other clinical features were studied. Adiponectin expression from isolated adipocytes was measured both in control and after insulin‐stimulation conditions by quantitative PCR.

Results  Subcutaneous adipocytes expressed significantly higher amounts of adiponectin mRNA than visceral tissue (P = 0·027). Insulin increased adiponectin expression specifically in the omental tissue (P = 0·011). In these patients, waist : hip ratio was directly correlated with adiponectin expression in the visceral depot (r = 0·660; P = 0·014), while fasting glucose levels were inversely associated with adiponectin mRNA in the subcutaneous tissue (r =–0·604; P = 0·022). Adiponectin expression after addition of insulin was positively correlated with some metabolic risk factors (cholesterol, LDL‐cholesterol, insulin, C‐peptide). Interestingly, local insulin induced an up‐regulation of adiponectin expression in the AT of those patients with higher metabolic syndrome disturbances.

Conclusions  Our results clearly demonstrate that insulin exerts a stimulating effect on adiponectin gene expression in a depot‐specific manner. The AT response to insulin stimulus depends on the physiopathological situation, being higher in those individuals with impaired insulin‐sensitivity and lipid metabolism.
Topographical associations between islet endocrine cells and duct epithelial cells in the adult human pancreas
Authors: Zhao Hai‐Lu , Sui Yi , Guan Jing , Lai Fernand M. M. , Gu Xue‐Mei , He Lan , Zhu Xun , Rowlands Dewi K. , Xu Gang , Tong Peter C. Y. , Chan Juliana C. N.
Published: August 2008

Objectives  The pancreatic ducts, endocrine islets and exocrine acini are three functionally related components. From birth to adulthood, the islets and ducts are regarded as independent entities. The objective of this study is to investigate the topographical associations between the islet endocrine cells and duct epithelial cells in the adult human pancreas.

Materials and methods  Panels of immunomarkers for the exocrine acinar cells (amylase), duct cells [cytokeratin 19 (CK19)], endocrine cells (chromogranin A, neurone specific enolase, synaptophysin) and islet hormones (glucagon, insulin, somatostatin, pancreatic polypeptide) were applied to sequential pancreatic tissue sections obtained from autopsy specimens of 10‐nondiabetic human adults. Double immunofluorescent staining with CK19 and islet hormones was performed to confirm the islet to duct interrelationship.

Results  Sequential sectioning and immunostaining showed that 45% of the 172 islets examined appeared as single endocrine cell units or small clusters of < 10 endocrine cells on at least one plane of section. A topographical association was found between the islet endocrine cells and duct epithelial cells. Topographical associations with CK 19‐stained duct cells occurred in 10·9% of the islet insulin‐containing β‐cells and in 8·9% of the islet glucagon‐producing α‐cells. The frequency of topographical associations increased toward the more distally located duct systems. The CK19‐stained duct cells and amylase‐labelled acinar cells were less frequently in association with other islet hormone‐producing cells.

Conclusions  Topographical associations between islet endocrine cells and pancreatic duct cells are frequent in adult human pancreas. The islet–duct association suggests possible functional interactions between the two interrelated pancreatic compartments.
Evaluation of the autoimmune regulator (AIRE) gene mutations in a cohort of Italian patients with autoimmune‐polyendocrinopathy‐candidiasis‐ectodermal‐dystrophy (APECED) and in their relatives
Authors: Cervato Sara , Mariniello Barbara , Lazzarotto Francesca , Morlin Luca , Zanchetta Renato , Radetti Giorgio , De Luca Filippo , Valenzise Mariella , Giordano Roberta , Rizzo Daniela , Giordano Carla , Betterle Corrado
Published: August 2008

Objective   Autoimmune‐polyendocrinopathy‐candidiasis‐ectodermal‐dystrophy (APECED) is a rare syndrome characterized by chronic candidiasis, chronic hypoparathyroidism and Addison's disease. APECED has been associated with mutations in autoimmune regulator (AIRE) gene. Our aim is to perform a genetic analysis of the AIRE gene in Italian APECED patients and in their relatives.

Design  AIRE mutations were determined by DNA sequencing in all subjects. Patients were tested for clinical autoimmune or non‐autoimmune diseases, or for organ and non‐organ specific autoantibodies.

Patients   A total of 24 Italian patients with APECED (15 from the Venetian region, 2 from Southern‐Tyrol, 4 from Apulia, 3 from Sicily), 25 relatives and 116 controls were studied.

Results  Ten out of the 15 Venetian patients (66%) were homozygous for R257X or compound heterozygous with 1094–1106del13. One patient was homozygous for 1094–1106del13 and another for R139X. A novel mutation (1032–1033delGT) in combination with 1094–1106del13 was identified in one patient. No mutations were found in two cases. Two patients from Southern Tyrol were homozygous for R257X and for 1094–1106del13bp. All patients from Apulia were homozygous or heterozygous for W78R combined with Q358X. The patients from Sicily were homozygous for R203X or compound heterozygous with R257X. The analysis of the genotype–phenotype revealed that patients carrying 1094–1106del13 at the onset of Addison's disease were significantly older than those carrying other mutations. The genetic study of 25 relatives identified 20 heterozygous subjects. They suffered from various autoimmune and non‐autoimmune diseases but no major disease of APECED was found.

Conclusion  These data demonstrate the great genetic heterogeneity for the AIRE mutations in Italian APECED patients, and that the heterozygosity for AIRE mutations do not produce APECED.