bluuebunny/crossref_metadata_2025 huggingface dataset:
Prognostic factors for patients with hepatocellular carcinoma with macroscopic portal vein or inferior vena cava tumor thrombi receiving external‐beam radiation therapy
Authors: Zeng Zhao‐Chong , Fan Jia , Tang Zhao‐You , Zhou Jian , Wang Jian‐Hua , Wang Bin‐Liang , Guo Wei
Published: November 2008

Prognostic factors in patients with hepatocellular carcinoma (HCC) with tumor thrombosis are not well established, especially for those given external‐beam radiation therapy (EBRT). Patients (n = 136) with HCC who had portal vein (PV) or inferior vena cava (IVC) tumor thrombus received EBRT between January 1998 and October 2007. Demographic variables, laboratory values, tumor characteristics, and treatment modalities were determined at diagnosis and before EBRT. The total radiation dose ranged from 30 to 60 Gy (median, 50 Gy) and was focused on the tumor thrombi. Predictors of survival were identified using the univariate and multivariate analysis. Of the 136 patients, the tumor thrombus completely disappeared in 41 patients (30.1%), 36 patients (26.5%) had a partial response, 49 patients (36%) had stable disease, and 10 patients (7.4%) had progressive disease. On multivariate analysis, pretreatment unfavorable predictors were associated with lower albumin, higher γ‐glutamyltransferase and α‐fetoprotein levels, poorer Child–Pugh classification, intrahepatic multifocality, lymph node metastases, poorer response to EBRT, and 2‐dimension EBRT technique. Survival rates at 1, 2, and 3 years were 31.8%, 17.5%, and 8.8% for patients with PV tumor thrombi; 66.3%, 21.1%, and 15.8% for IVC tumor thrombi; and 25%, 8.3%, and 0% for PV plus IVC tumor thrombi, respectively. Overall median survival was 9.7 months. This study provides detailed information about the survival outcomes and prognostic factors of HCC with tumor thrombi in a relatively large cohort of patients treated with radiation, and the results will help in understanding the potential factors that influence survival for patients with HCC after EBRT. (Cancer Sci 2008; 99: 2510–2517)
Suppression of transient receptor potential melastatin 7 channel induces cell death in gastric cancer
Authors: Kim Byung Joo , Park Eun Jung , Lee Jae Hwa , Jeon Ju‐Hong , Kim Seon Jeong , So Insuk
Published: November 2008

Ca2+ and Mg2+ have a fundamental role in many cellular processes and ion channels are involved in normal physiologic processes and in the pathology of various diseases. The aim here was to show that the presence and potential role of transient receptor potential melastatin 7 (TRPM7) channels in the growth and survival of AGS cells, the most common human gastric adenocarcinoma cell line. The patch‐clamp technique for whole‐cell recording was used in AGS cells. TRPM7‐specific small interfering RNAs were used for specific inhibition of TRPM7. Whole‐cell voltage‐clamp recordings revealed the TRPM7‐like currents that activated spontaneously following loss of intracellular Mg2+. The current had a non‐linear current–voltage relationship with the characteristic steep outward rectification associated with TRPM7 channels. Reverse transcription–polymerase chain reaction, western blotting, and immunoreactivity all showed abundant expression of TRPM7 messenger RNA and protein in AGS cells. Transfection of AGS cells with TRPM7 siRNA significantly reduced the expression of TRPM7 mRNA and protein as well as the amplitude of the TRPM7‐like currents. Furthermore, we found that Mg2+ is critical for the growth and survival in AGS cells. Blockade of TRPM7 channels by La3+ and 2‐APB or suppression of TRPM7 expression by siRNA inhibited the growth and survival of these cells. Human gastric adenocarcinoma cells express TRPM7 channel whose presence is essential for cell survival. The protein is a likely potential target for the pharmacological treatment of gastric cancer. (Cancer Sci 2008; 99: 2502–2509)
Roles of cyclooxygenase‐2 and microsomal prostaglandin E synthase‐1 expression and β‐catenin activation in gastric carcinogenesis inN‐methyl‐N‐nitrosourea‐treated K19‐C2mE transgenic mice
Authors: Takasu Shinji , Tsukamoto Tetsuya , Cao Xue‐Yuan , Toyoda Takeshi , Hirata Akihiro , Ban Hisayo , Yamamoto Masami , Sakai Hiroki , Yanai Tokuma , Masegi Toshiaki , Oshima Masanobu , Tatematsu Masae
Published: November 2008

K19‐C2mE transgenic (Tg) mice, simultaneously expressing cyclooxygenase‐2 (COX‐2) and microsomal prostaglandin E synthase‐1 (mPGES‐1) in the gastric mucosa under the cytokeratin 19 gene promoter, were here treated withN‐methyl‐N‐nitrosourea (MNU) and inoculated withHelicobacter pylori(H. pylori) to investigate gastric carcinogenesis. Wild‐type (WT) and Tg mice undergoing MNU treatment frequently developed tumors in the pyloric region (100% and 94.7%, respectively); multiplicity in Tg was higher than that in WT (P < 0.05) withH. pyloriinfection. Larger pyloric tumors were more frequently observed in Tg than in WT (P < 0.05). In addition, Tg developed fundic tumors, where WT did not. No gastric tumors were observed without MNU treatment. Transcripts of TNF‐α, iNOS, IL‐1β, and CXCL14 were up‐regulated withH. pyloriinfection in both genotypes and were also increased more in Tg than in WT withinH. pylori‐inoculated animals. Immunohistochemical analysis demonstrated significantly greater β‐catenin accumulation in pyloric tumors, compared with those in the fundus (P < 0.01) with mutations of exon 3; 18.2% and 31.6% in MNU‐alone and MNU + H. pylori‐treated WT, whereas 21.4% and 62.5% was observed in the Tg, respectively; the latter significantly higher (P < 0.05), suggesting the role ofH. pyloriin Wnt activation. In conclusion, K19‐C2mE mice promoted gastric cancer in both fundic and pyloric regions. Furthermore β‐catenin activation may play the important role of pyloric carcinogenesis especially inH. pylori‐infected Tg. Induction of various inflammatory cytokines in addition to overexpression of COX‐2/mPGES‐1 could be risk factors of gastric carcinogenesis and may serve as a better gastric carcinogenesis model. (Cancer Sci2008; 99: 2356–2364)
Low‐intensity ultrasound and microbubbles enhance the antitumor effect of cisplatin
Authors: Watanabe Yukiko , Aoi Atsuko , Horie Sachiko , Tomita Noriko , Mori Shiro , Morikawa Hidehiro , Matsumura Yasuhiro , Vassaux Georges , Kodama Tetsuya
Published: November 2008

Cell permeabilization using microbubbles (MB) and low‐intensity ultrasound (US) have the potential for delivering molecules into the cytoplasm. The collapsing MB and cavitation bubbles created by this collapse generate impulsive pressures that cause transient membrane permeability, allowing exogenous molecules to enter the cells. To evaluate this methodology in vitro and in vivo, we investigated the effects of low‐intensity 1‐MHz pulsed US and MB combined with cis‐diamminedichloroplatinum (II) (CDDP) on two cell lines (Colon 26 murine colon carcinoma and EMT6 murine mammary carcinoma) in vitro and in vivo on severe combined immunodeficient mice inoculated with HT29‐luc human colon carcinoma. To investigate in vitro the efficiency of molecular delivery by the US and MB method, calcein molecules with a molecular weight in the same range as that of CDDP were used as fluorescent markers. Fluorescence measurement revealed that approximately 106–107 calcein molecules per cell were internalized. US–MB‐mediated delivery of CDDP in Colon 26 and EMT6 cells increased cytotoxicity in a dose‐dependent manner and induced apoptosis (nuclear condensation and fragmentation, and increase in caspase‐3 activity). In vivo experiments with xenografts (HT29‐luc) revealed a very significant reduction in tumor volume in mice treated with CDDP + US + MB compared with those in the US + CDDP groups for two different concentrations of CDDP. This finding suggests that the US–MB method combined with chemotherapy has clinical potential in cancer therapy. (Cancer Sci 2008; 99: 2525–2531)
Inhibition of α‐mannosidase Man2c1 gene expression suppresses growth of esophageal carcinoma cells through mitotic arrest and apoptosis
Authors: Tian Yun , Ju Ji Yu , Zhou Yi Qun , Liu Yin , Zhu Li Ping
Published: November 2008

To study the effects of suppressed α‐mannosidase Man2c1 gene expression on EC9706 human esophageal carcinoma cells, the cells were treated with short interfering RNA. Growth inhibition of EC9706 cells was observed when Man2c1 expression was inhibited in this way. Flow cytometric analysis showed accumulation of cells in S and G2–M phases, as well as cell apoptosis. The mitotic index test showed cell‐cycle arrest at the M checkpoint. Although the percentage of cells in (pro)metaphase increased, the proportion of cells in anaphase and telophase decreased. Apoptosis was trigged by mitotic arrest. Furthermore, microtubules in EC9607 cells were examined by means of fluorescence staining of α‐tubulin. Although control cells showed a nest‐like microtubule network, the microtubule network in experimental cells was vague and condensed at the perinuclear region. Some cells with Man2c1 suppression had large protrusions of cytoplasm, some of which linked with the main body through a long, thin connection. Western blotting showed that tubulin polymerization was inhibited. The data imply that induction of mitotic arrest and consequent apoptosis resulted from microtubule disorganization, which appears to be one of the major cellular mechanisms by which suppressed expression of the Man2c1 gene causes growth inhibition of EC9706 esophageal carcinoma cells. In addition, Man2c1 suppression results in upregulation of E‐cadherin, α‐catenin, and β‐catenin expression in cells. (Cancer Sci 2008; 99: 2428–2434)
The social context of parenting 3‐year‐old children with developmental delay in the UK
Authors: Emerson E. , Graham H. , McCulloch A. , Blacher J. , Hatton C. , Llewellyn G.
Published: November 2008

Background  Children with intellectual or developmental disability have significantly poorer health and mental health than their non‐disabled peers and are at high risk of social exclusion. The aim of the present paper is to provide information on the circumstances in which 3‐year‐old children at risk of intellectual or developmental disability are growing up in the UK.

Methods  Secondary analysis of data on 12 689 families in English‐speaking monolingual households from the first two waves of the UK's Millennium Cohort Study. A total of 440 children (3% of the weighted sample) were identified as being developmentally delayed.

Results  When compared with other children, children with developmental delays were more disadvantaged on every indicator of social and economic disadvantage examined. Two out of three children with developmental delays had been exposed to repeated disadvantage as measured by income poverty, material hardship, social housing and receipt of means‐tested benefits. The effect of repeated disadvantage on the risk of developmental delay remained after account was taken of parental education and occupational status.

Conclusions  Young children with delayed development in the UK are likely to be exposed to repeated socio‐economic disadvantage. Implications for policy and understanding the nature of the link between poverty and child disability are discussed.
Is aggression in children with behavioural and emotional difficulties associated with television viewing and video game playing? A systematic review
Authors: Mitrofan O. , Paul M. , Spencer N.
Published: November 2008

Background  Possible associations between television viewing and video game playing and children's aggression have become public health concerns. We did a systematic review of studies that examined such associations, focussing on children and young people with behavioural and emotional difficulties, who are thought to be more susceptible.

Methods  We did computer‐assisted searches of health and social science databases, gateways, publications from relevant organizations and for grey literature; scanned bibliographies; hand‐searched key journals; and corresponded with authors. We critically appraised all studies.

Results  A total of 12 studies: three experiments with children with behavioural and emotional difficulties found increased aggression after watching aggressive as opposed to low‐aggressive content television programmes, one found the opposite and two no clear effect, one found such children no more likely than controls to imitate aggressive television characters. One case–control study and one survey found that children and young people with behavioural and emotional difficulties watched more television than controls; another did not. Two studies found that children and young people with behavioural and emotional difficulties viewed more hours of aggressive television programmes than controls. One study on video game use found that young people with behavioural and emotional difficulties viewed more minutes of violence and played longer than controls. In a qualitative study children with behavioural and emotional difficulties, but not their parents, did not associate watching television with aggression. All studies had significant methodological flaws. None was based on power calculations.

Conclusion  This systematic review found insufficient, contradictory and methodologically flawed evidence on the association between television viewing and video game playing and aggression in children and young people with behavioural and emotional difficulties. If public health advice is to be evidence‐based, good quality research is needed.
The volatile anaesthetic sevoflurane attenuates lipopolysaccharide-induced injury in alveolar macrophages
Authors: Steurer M , Schläpfer M , Steurer M , Z'graggen B Roth , Booy C , Reyes L , Spahn D R , Beck-Schimmer B
Published: November 2008

Acute lung injury (ALI) is a well-defined inflammation whereby alveolar macrophages play a crucial role as effector cells. As shown previously in numerous experimental approaches, volatile anaesthetics might reduce the degree of injury in pre- or post-conditioning set-ups. Therefore, we were interested to evaluate the effect of the application of the volatile anaesthetic sevoflurane on alveolar macrophages regarding the expression of inflammatory mediators upon lipopolysaccharide (LPS) stimulation in vitro. Alveolar macrophages were stimulated with LPS. Two hours later, cells were exposed additionally to air (control) or to sevoflurane-containing air for 4, 6, 8, 12 or 24 h. Tumour necrosis factor (TNF)-α, cytokine-induced neutrophil chemoattractant-1 (CINC-1), macrophage-inflammatory protein-2 (MIP-2) and monocyte chemoattractant protein-1 (MCP-1) proteins were determined and chemotaxis assays were performed. To evaluate possible cellular signalling pathways phosphorylation of the kinases extracellular-regulated kinase (ERK) and Akt was assessed. In the early phase of sevoflurane post-conditioning expression of TNF-α, CINC-1, MIP-2 and MCP-1 was attenuated, leading to a diminished chemotaxis reaction for neutrophils. Phosphorylation of ERK seems to be a possible cellular mechanism in the sevoflurane-induced protection in vitro. Pharmacological post-conditioning of alveolar macrophages with sevoflurane immunmodulates the inflammatory response upon stimulation with endotoxin. This might be a possible option for a therapeutical approach in ALI.
Climate, oceanography, and recruitment: the case of the Bay of Biscay anchovy (Engraulis encrasicolus)
Authors: BORJA ANGEL , FONTÁN ALMUDENA , SÁENZ JON , VALENCIA VICTORIANO
Published: November 2008

The Bay of Biscay anchovy has experienced, since 2001, a succession of low recruitments, resulting in the collapse of the stock in 2005; this has led to successive closures of the fishery. This study investigates the possible impact of different controlling factors [North Atlantic Oscillation (NAO), East Atlantic (EA) pattern, turbulence, upwelling, and river flow] upon anchovy recruitment and fishery catches. Fifty‐five percent of the recruitment variability of this fishery can be explained by upwelling over the spawning area; this is related, in turn, to the EA pattern. The conceptual understanding of the system proposed for the Bay of Biscay anchovy suggests that negative EA periods are associated with northeasterly wind circulation, which produces weak upwelling over the continental shelf. This pattern results in hydrodynamic stability over the area, leading, probably, to adequate food availability. A positive EA (which extends onwards, from 1998) is associated with southwesterly winds and downwelling over the continental shelf; this leads, probably, to the dispersion of anchovy food and larvae, together with increasing mortality.
Cell adhesion markedly increases lucigenin‐enhanced chemiluminescence of the phagocyte NADPH oxidase
Authors: Kuribayashi Futoshi , Tsuruta Satoru , Yamazaki Tsuyoshi , Nunoi Hiroyuki , Imajoh‐Ohmi Shinobu , Kanegasaki Shiro , Nakamura Michio
Published: November 2008

Lucigenin‐enhanced chemiluminescence (LECL) is widely used for the detection of reactive oxygen species released from various cells and mitochondria. However, the LECL response varies depending on cell species and assay conditions at least in part by unknown factors. Here we report that cell adhesion is an important factor for increasing LECL of tetradecanoylphorbol acetate (TPA)‐stimulated human neutrophils. More than 90% LECL remained even after complete removal of the cell suspension 10 min after TPA stimulation, and ~22.5% of neutrophils were adhered to the reaction tube. These results indicate that LECL by an adhering neutrophil is ~45× higher than that by a non‐adhering neutrophil. LECL by leukocyte adhesion deficiency neutrophils was one‐fifth of that by normal neutrophils and completely disappeared when the cell suspension was removed, confirming that LECL depends highly on cell adhesion. The oxidase activity of adhering neutrophils measured after permeabilization with Renex 30 together with NADPH addition was similar to that of non‐adhering neutrophils, indicating that lucigenin and cell adhesion do not enhance the oxidase activity. Based on these findings, we propose that a mixture of adhering and non‐adhering neutrophils can be used for simultaneous screenings of adhering activity and the oxidase activity of neutrophils.
Effects of Akt signaling on nuclear reprogramming
Authors: Nakamura Toshinobu , Inoue Kimiko , Ogawa Shoko , Umehara Hiroki , Ogonuki Narumi , Miki Hiromi , Kimura Tohru , Ogura Atsuo , Nakano Toru
Published: November 2008

Reprogramming of the epigenetic state from differentiated to pluripotent cells can be attained by cell fusion of differentiated somatic cells with embryonic stem (ES) cells or transfer of the nucleus of a differentiated cell into an enucleated oocyte. Activation of Akt signaling is sufficient to maintain pluripotency of ES cells and promotes derivation of embryonic germ (EG) cells from primordial germ cells (PGCs). Here we analyzed the effects of Akt signaling on somatic cell nuclear reprogramming after cell fusion and nuclear transfer. We found that forced activation of Akt signaling stimulated reprogramming after cell fusion of ES cells with thymocytes or mouse embryonic fibroblasts. These hybrid cells showed ES cell characteristics, including in vitro and in vivo differentiation capacity. In contrast, Akt signaling significantly reduced the efficiency of reprogramming with nuclear transfer. Our results demonstrate that Akt signaling plays important roles on the nuclear reprogramming of somatic cells.
c‐Maf increases apoptosis in peripheral CD8 cells by transactivating Caspase 6
Authors: Peng Siying , Wu Hailong , Mo Yin‐Yuan , Watabe Kounosuke , Pauza Mary E.
Published: November 2008

In addition to transactivation of interleukin‐4 (IL‐4), cellular muscular aponeurotic fibrosarcoma (c‐Maf) enhances CD4 cell apoptosis by limiting Bcl‐2 expression. The CD8 cells also express c‐Maf and peripheral CD8 cell numbers are reduced in c‐Maf transgenic mice, suggesting that c‐Maf may influence CD8 cell survival in a manner similar to CD4 cells. Here we confirm that, similar to CD4 cells, c‐Maf enhances CD8 cell susceptibility to apoptosis induced by multiple stimuli, independent of IL‐4. However, unlike CD4 cells, c‐Maf enhancement of apoptosis is independent of Bcl‐2, suggesting that c‐Maf uses other mechanisms to regulate CD8 cell apoptosis. Real‐time reverse transcription–polymerase chain reaction reveals that the pro‐apoptotic gene Caspase 6 is upregulated in c‐Maf transgenic CD8 cells, suggesting that Caspase 6 is a novel c‐Maf target gene. Luciferase reporter assays and site‐directed mutagenesis reveal a functional c‐Maf recognition element (MARE) within the first intron of Caspase 6. Binding of c‐Maf to the MARE site is detectable by chromatin immunoprecipitation using non‐transgenic T‐cell lysates, so c‐Maf can interact with the Caspase 6 MARE site in normal T cells. Furthermore, caspase 6 activity is increased among CD8 cells from c‐Maf transgenic mice following T‐cell receptor engagement. As expected, activity of the downstream caspases 3 and 7 is also increased. Consistent with the ability of caspase 6 to participate in positive feedback loops, cytochrome c release and caspase 8 activation are also increased. Together these results indicated that c‐Maf increases CD8 cell sensitivity to apoptotic stimuli, at least in part, by direct transactivation of Caspase 6, providing increased substrate for Caspase 6‐dependent apoptosis pathways.
Mannan‐binding lectin deficiency modulates the humoral immune response dependent on the genetic environment
Authors: Ruseva Marieta , Kolev Martin , Dagnaes‐Hansen Frederik , Hansen Soren B. , Takahashi Kazue , Ezekowitz Alan , Thiel Steffen , Jensenius Jens C. , Gadjeva Mihaela
Published: November 2008

Mannan‐binding lectin (MBL) is a plasma protein implicated in innate immune defence against a broad range of microorganisms, including viruses. It is also thought that MBL plays a role in the recruitment of the specific clonal immune response. This was studied by injecting soluble hepatitis B surface antigen (HBsAg) intravenously into mice deficient in both MBL‐A and MBL‐C (MBL DKO mice). The MBL DKO animals on mixed genetic background (SV129EvSv × C57BL/6) produced higher antibody titres than the wild‐type littermates. After primary challenge with the antigen the immunoglobulin M anti‐HBsAg antibody titres were threefold higher in the MBL DKO mice than in the wild‐type mice. Following the boost, the immunoglobulin G anti‐HBsAg antibody titres were 10‐fold higher in the MBL DKO mice, suggesting that MBL plays a role in a negative feedback regulation of adaptive immunity. However, the modulating effect of MBL was dependent on the genetic environment. The MBL DKO mice backcrossed on a C57BL/6 background showed the opposite response with the MBL DKO mice now producing fewer antibodies than the wild‐type animals, whereas MBL deficiency in mice with the SV129EvSv background did not show any effect in antibody production. These findings indicate that the modifying effect of MBL on the humoral immune response is influenced by the genetic environment.
Interleukin‐17 in host defence against bacterial, mycobacterial and fungal pathogens
Authors: Curtis Meredith M. , Way Sing Sing
Published: November 2008

The mammalian immune system is intricately regulated, allowing for potent pathogen‐specific immunity to be rapidly activated in response to infection with a broad and diverse array of potential pathogens. As a result of their ability to differentiate into distinct effector lineages, CD4 T cells significantly contribute to pathogen‐specific adaptive immune responses. Through the production of effector cytokines, CD4 T helper (Th) cells orchestrate the precise mobilization of specific immune cells to eradicate infection. The protective effects of the newly identified lineage of Th17 cells against pathogens like Klebsiella pneumoniae, Citrobacter rodentium and Candida albicans indicate the capacity of Th17 cells to confer protection against extracellular bacterial and fungal pathogens, filling a critical void in host immunity not covered by the classically described Th1 lineage that activates immunity to intracellular pathogens or the Th2 lineage that is important in protection against mucosal parasitic pathogens. Host defence by Th17 cells extends beyond protection against extracellular bacterial and fungal pathogens, as demonstrated in infections against intracellular bacteria like Listeria monocytogenes and Salmonella enterica, as well as Mycobacterium tuberculosis. Herein, we summarize both experimental data from mouse infection models and epidemiological studies in humans that demonstrate the protective effects of interleukin‐17 and Th17 CD4 T cells in immunity to bacterial, mycobacterial and fungal pathogens.
Habitat area but not habitat age determines wild bee richness in limestone quarries
Authors: Krauss Jochen , Alfert Thomas , Steffan‐Dewenter Ingolf
Published: November 2008

Within highly modified European landscapes, limestone quarries can act as important secondary habitats for a range of endangered wild bee species. However, the relative influence of quarry habitat area, habitat age and within‐habitat diversity on the conservation value of these secondary habitats is mainly unknown.

We assessed species richness and abundance of wild bees by variable transect walks in 24 limestone quarries ranging in size from 0·01 to 21·2 ha. Species traits such as social status, resource specialization and nesting substrate were used to define functional guilds of bees.

In total, 41% of all wild bee species known from southern Lower Saxony, Germany, were found in the studied quarries. Total species richness increased with habitat area but, in contrast to our expectations, not with habitat age, although we tested an age gradient of over 120 years. Solitary species were more strongly affected by decreasing habitat area than social species but response did not differ with respect to habitat age.

Hierarchical partitioning analyses revealed that habitat area per se, within‐habitat diversity and sampling effort were of similar importance in explaining species richness patterns.

Synthesis and applications. Even newly created limestone quarries provide an important secondary habitat for wild bees. Therefore, maintenance and management of secondary succession of these sites should be given high priority in species conservation plans. Landscape management schemes involving filling or flooding such quarries should be prevented.
Genetic and acoustic population structuring in the Okinawa least horseshoe bat: are intercolony acoustic differences maintained by vertical maternal transmission?
Authors: YOSHINO HAJIME , ARMSTRONG KYLE N. , IZAWA MASAKO , YOKOYAMA JUN , KAWATA MASAKADO
Published: November 2008

The origin and meaning of echolocation call frequency variation within rhinolophid bats is not well understood despite an increasing number of allopatric and sympatric examples being documented. A bimodal distribution of mean regional call frequency within the Okinawa‐jima Island population of Rhinolophus cornutus pumilus (Rhinolophidae) provided a unique opportunity to investigate geographic call frequency variation early in its development. Individual resting echolocation frequencies, partial mitochondrial DNA D‐loop sequences and genotypes from six microsatellite loci were obtained from 288 individuals in 11 colonies across the entire length of the island, and nearby Kume‐jima Island. Acoustic differences (5–8 kHz) observed between the north and south regions have been maintained despite evidence of sufficient nuclear gene flow across the middle of the island. Significant subdivision of maternally inherited D‐loop haplotypes suggested a limitation of movement of females between regions, but not within the regions, and was evidence of female philopatry. These results support a ‘maternal transmission’ hypothesis whereby the difference in the constant frequency (CF) component between the regions is maintained by mother–offspring transmission of CF, the restricted dispersal of females between regions and small effective population size. We suggest that the mean 5–8 kHz call frequency difference between the regions might develop through random cultural drift.
Herbivore‐induced jasmonic acid bursts in leaves of Nicotiana attenuata mediate short‐term reductions in root growth
Authors: HUMMEL GRÉGOIRE M. , SCHURR ULRICH , BALDWIN IAN T. , WALTER ACHIM
Published: November 2008

Root growth in Nicotiana attenuata is transiently reduced after application of oral secretions (OS) of Manduca sexta larvae to wounds in leaves. Feeding of M. sexta or OS elicitation is known to result in jasmonic acid (JA) and ethylene bursts, and activates a suite of defence responses. Because both plant hormones are known to strongly reduce root growth, their activation might account for the observed reduction of root growth following herbivory. To test this hypothesis, we measured primary root growth with digital image sequence processing at high temporal resolution in antisense‐lipoxygenase 3 (asLOX3) and inverted repeat‐coronatin‐insensitive 1 (irCOI1) seedlings which are impaired in JA biosynthesis and perception, respectively, and wild‐type (WT) seedlings. Higher root growth rates in irCOI1 compared with WT were observed after OS elicitation. The dynamics of wound‐induced root growth reduction coincide with the dynamics of root growth reduction induced by external application of methyl JA. In an experiment with 1‐methylcyclopropen (1‐MCP), a potent ethylene receptor blocker, no wounding‐specific difference between growth of 1‐MCP‐treated plants and non‐treated plants was observed, suggesting that wound‐induced endogenous JA and not ethylene mediates the wounding‐specific reduction in root growth. Yet, inhibiting the ethylene response by applying 1‐MCP led to markedly increased root growth compared with that of control plants, indicating that ethylene normally suppresses plant growth in N. attenuata seedlings.
The historical record in the Scaglia limestone at Gubbio: magnetic reversals and the Cretaceous‐Tertiary mass extinction
Authors: ALVAREZ WALTER
Published: November 2008

The Scaglia limestone in the Umbria‐Marche Apennines, well‐exposed in the Gubbio area, offered an unusual opportunity to stratigraphers. It is a deep‐water limestone carrying an unparalleled historical record of the Late Cretaceous and Palaeogene, undisturbed by erosional gaps. The Scaglia is a pelagic sediment largely composed of calcareous plankton (calcareous nannofossils and planktonic foraminifera), the best available tool for dating and long‐distance correlation. In the 1970s it was recognized that these pelagic limestones carry a record of the reversals of the magnetic field. Abundant planktonic foraminifera made it possible to date the reversals from 80 to 50 Ma, and subsequent studies of related pelagic limestones allowed the micropalaeontological calibration of more than 100 Myr of geomagnetic polarity stratigraphy, from ca 137 to ca 23 Ma. Some parts of the section also contain datable volcanic ash layers, allowing numerical age calibration of the reversal and micropalaeontological time scales. The reversal sequence determined from the Italian pelagic limestones was used to date the marine magnetic anomaly sequence, thus putting ages on the reconstructed maps of continental positions since the breakup of Pangaea. The Gubbio Scaglia also contains an apparently continuous record across the Cretaceous–Tertiary boundary, which was thought in the 1970s to be marked everywhere in the world by a hiatus. A geochemical study of the Gubbio Cretaceous–Tertiary boundary accidentally uncovered an unexpectedly large concentration of iridium in the boundary clay, providing the first evidence for a large comet or asteroid impact at the time of the Cretaceous–Tertiary mass extinction. The sudden foraminiferal extinction in the essentially complete Gubbio Scaglia showed that the Cretaceous–Tertiary extinction was a catastrophic event. After publication of the Gubbio Cretaceous–Tertiary iridium anomaly in 1980, a decade followed in which more and more evidence for a catastrophic impact challenged the traditional uniformitarian viewpoint of geologists. In 1991, the impact site was discovered at Chicxulub, in the Yucatán Peninsula of Mexico. The Gubbio area has thus emerged as a classical geological locality. It has been critical in the development of geomagnetic reversal stratigraphy, a third system for dating Earth history, complementing biostratigraphy and radiometric age dating. Gubbio was also central to the recognition that occasional catastrophic events like great impacts require a rejection of strict uniformitarianism.
Quantification of allele‐specific expression of a gene encoding strawberry polygalacturonase‐inhibiting protein (PGIP) using PyrosequencingTM
Authors: Schaart Jan G. , Mehli Lisbeth , Schouten Henk J.
Published: November 2008

Recent studies indicate that allele‐specific differences in gene expression are a common phenomenon. The extent to which differential allelic expression exists might be underestimated, due to the limited accuracy of the methods used so far. To demonstrate allele‐specific expression, we investigated the transcript abundance of six individual, highly homologous alleles of a polygalacturonase‐inhibiting protein gene (FaPGIP) from octoploid strawberry (Fragaria × ananassa). We applied the highly quantitative Pyrosequencing method which, for the gene under study, detected allele frequency differences as small as 4.0 ± 2.8%. Pyrosequencing of RT‐PCR products showed that one FaPGIP allele was preferentially expressed in leaf tissue, while two other alleles were expressed in a fruit‐specific way. For fruits that were inoculated with Botrytis cinerea a strong increase in overall FaPGIP gene expression was observed. This upregulation was accompanied by a significant change in FaPGIP allele frequencies when compared with non‐treated fruits. Remarkably, in the five cultivars tested, the allele frequency in cDNA from the inoculated fruits was similar to that in genomic DNA, suggesting uniform upregulation of all FaPGIP alleles present as a result of pathogenesis‐related stress. The results demonstrate that when Pyrosequencing of RT‐PCR products is performed, novel allele‐specific gene regulation can be detected and quantified.
HUA2 is required for the expression of floral repressors in Arabidopsis thaliana
Authors: Doyle Mark R. , Bizzell Colleen M. , Keller Melissa R. , Michaels Scott D. , Song Judong , Noh Yoo‐Sun , Amasino Richard M.
Published: November 2008

The HUA2 gene acts as a repressor of floral transition. Lesions in hua2 were identified through a study of natural variation and through two mutant screens. An allele of HUA2 from Landsberg erecta (Ler) contains a premature stop codon and acts as an enhancer of early flowering 4 (elf4) mutants. hua2 single mutants, in the absence of the elf4 lesion, flower earlier than wild type under short days. hua2 mutations partially suppress late flowering in FRIGIDA (FRI )‐containing lines, autonomous pathway mutants, and a photoperiod pathway mutant. hua2 mutations suppress late flowering by reducing the expression of several MADS genes that act as floral repressors including FLOWERING LOCUS C (FLC ) and FLOWERING LOCUS M (FLM ).
Molecular and functional characterization of Arabidopsis Cullin 3A
Authors: Dieterle Monika , Thomann Alexis , Renou Jean‐Pierre , Parmentier Yves , Cognat Valerie , Lemonnier Gaetan , Müller Rebecca , Shen Wen‐Hui , Kretsch Thomas , Genschik Pascal
Published: November 2008

Cullin proteins, which belong to multigenic families in all eukaryotes, associate with other proteins to form ubiquitin protein ligases (E3s) that target substrates for proteolysis by the 26S proteasome. Here, we present the molecular and genetic characterization of a plant Cullin3. In contrast to fungi and animals, the genome of the model plant Arabidopsis thaliana contains two related CUL3 genes, called CUL3A and CUL3B. We found that CUL3A is ubiquitously expressed in plants and is able to interact with the ring‐finger protein RBX1. A genomic search revealed the existence of at least 76 BTB‐domain proteins in Arabidopsis belonging to 11 major families. Yeast two‐hybrid experiments indicate that representative members of certain families are able to physically interact with both CUL3A and CUL3B, suggesting that Arabidopsis CUL3 forms E3 protein complexes with certain BTB domain proteins. In order to determine the function of CUL3A, we used a reverse genetic approach. The cul3a null mutant flowers slightly later than the control plants. Furthermore, this mutant exhibits a reduced sensitivity of the inhibition of hypocotyl growth in far‐red light and miss‐expresses COP1. The viability of the mutant plants suggests functional redundancy between the two CUL3 genes in Arabidopsis.
ALS3 encodes a phloem‐localized ABC transporter‐like protein that is required for aluminum tolerance in Arabidopsis
Authors: Larsen Paul B. , Geisler Matt J.B. , Jones Carol A. , Williams Kelly M. , Cancel Jesse D.
Published: November 2008

Aluminum (Al) toxicity in acid soils is a worldwide agricultural problem that severely limits crop productivity through inhibition of root growth. Previously, Arabidopsis mutants with increased Al sensitivity were isolated in order to identify genes important for Al tolerance in plants. One mutant, als3, exhibited extreme root growth inhibition in the presence of Al, suggesting that this mutation negatively impacts a gene required for Al tolerance. Map‐based cloning of the als3‐1 mutation resulted in the isolation of a novel gene that encodes a previously undescribed ABC transporter‐like protein, which is highly homologous to a putative bacterial metal resistance protein, ybbM. Northern analysis for ALS3 expression revealed that it is found in all organs examined, which is consistent with the global nature of Al sensitivity displayed by als3, and that expression increases in roots following Al treatment. Based on GUS fusion and in situ hybridization analyses, ALS3 is primarily expressed in leaf hydathodes and the phloem throughout the plant, along with the root cortex following Al treatment. Immunolocalization indicates that ALS3 predominantly accumulates in the plasma membrane of cells that express ALS3. From our results, it appears that ALS3 encodes an ABC transporter‐like protein that is required for Al resistance/tolerance and may function to redistribute accumulated Al away from sensitive tissues in order to protect the growing root from the toxic effects of Al.
In vivo studies on the roles of Tic110, Tic40 and Hsp93 during chloroplast protein import
Authors: Kovacheva Sabina , Bédard Jocelyn , Patel Ramesh , Dudley Penny , Twell David , Ríos Gabino , Koncz Csaba , Jarvis Paul
Published: November 2008

A multisubunit translocon of the inner envelope membrane, termed Tic, mediates the late stages of protein import into chloroplasts. Membrane proteins, Tic110 and Tic40, and a stromal chaperone, Hsp93, have been proposed to function together within the Tic complex. In Arabidopsis, single genes, atTIC110 and atTIC40, encode the Tic proteins, and two homologous genes, atHSP93‐V and atHSP93‐III, encode Hsp93. These four genes exhibited relatively uniform patterns of expression, suggesting important roles for plastid biogenesis throughout development and in all tissues. To investigate the roles played by these proteins in vivo, we conducted a comparative study of T‐DNA knockout mutants for each Tic gene, and for the most abundantly expressed Hsp93 gene, atHSP93‐V. In the homozygous state, the tic110 mutation caused embryo lethality, implying an essential role for atTic110 during plastid biogenesis. Homozygous tic110 embryos exhibited retarded growth, developmental arrest at the globular stage and a ‘raspberry‐like’ embryo‐proper phenotype. Heterozygous tic110 plants, and plants homozygous for the tic40 and hsp93‐V mutations, exhibited chlorosis, aberrant chloroplast biogenesis, and inefficient chloroplast‐import of both photosynthetic and non‐photosynthetic preproteins. Non‐additive interactions amongst the mutations occurred in double mutants, suggesting that the three components may cooperate during chloroplast protein import.
A study in scarlet: enzymes of ketocarotenoid biosynthesis in the flowers of Adonis aestivalis
Authors: Cunningham Francis X. , Gantt Elisabeth
Published: November 2008

The red ketocarotenoid astaxanthin (3,3′‐dihydroxy‐4,4′‐diketo‐β,β‐carotene) is widely used as an additive in feed for the pigmentation of fish and crustaceans and is frequently included in human nutritional supplements as well. There is considerable interest in developing a plant‐based biological production process for this valuable carotenoid. Adonis aestivalis (Ranunculaceae) is unusual among plants in synthesizing and accumulating large amounts of astaxanthin and other ketocarotenoids. The formation of astaxanthin requires only the addition of a carbonyl at the number 4 carbon of each β‐ring of zeaxanthin (3,3′‐dihydroxy‐β,β‐carotene), a carotenoid typically present in the green tissues of higher plants. We screened an A. aestivalis flower library to identify cDNAs that might encode the enzyme that catalyzes the addition of the carbonyls. Two closely related cDNAs selected in this screen were found to specify polypeptides similar in sequence to plant β‐carotene 3‐hydroxylases, enzymes that convert β‐carotene (β,β‐carotene) into zeaxanthin. The Adonis enzymes, however, exhibited neither 4‐ketolase nor 3‐hydroxylase activity when presented with β‐carotene as the substrate in Escherichia coli. Instead, the products of the Adonis cDNAs were found to modify β‐rings in two distinctly different ways: desaturation at the 3,4 position and hydroxylation of the number 4 carbon. The 4‐hydroxylated carotenoids formed in E. coli were slowly metabolized to yield compounds with ketocarotenoid‐like absorption spectra. It is proposed that a 3,4‐desaturation subsequent to 4‐hydroxylation of the β‐ring leads to the formation of a 4‐keto‐β‐ring via an indirect and unexpected route: a keto‐enol tautomerization.
Arabidopsis SHMT1, a serine hydroxymethyltransferase that functions in the photorespiratory pathway influences resistance to biotic and abiotic stress
Authors: Moreno Juan Ignacio , Martín Raquel , Castresana Carmen
Published: November 2008

We found that a recessive mutation, shmt1‐1, causes aberrant regulation of cell death resulting in chlorotic and necrotic lesion formation under a variety of environmental conditions. Salicylic acid‐inducible genes and genes involved in H2O2 detoxification were expressed constitutively in shmt1‐1 plants in direct correlation with the severity of the lesions. The shmt1‐1 mutants were more susceptible than control plants to infection with biotrophic and necrotrophic pathogens, developing severe infection symptoms in a high percentage of infected leaves. In addition, mutants carrying shmt1‐1 or a loss‐of‐function shmt1‐2 allele, were smaller and showed a greater loss of chlorophyll and greater accumulation of H2O2 than wild‐type plants when subjected to salt stress. SHMT1 was map‐based cloned and found to encode a serine hydroxymetyltransferase (SHMT1) involved in the photorespiratory pathway. Our results indicate that this enzymatic activity plays a critical role in controlling the cell damage provoked by abiotic stresses such as high light and salt and in restricting pathogen‐induced cell death, supporting the notion that photorespiration forms part of the dissipatory mechanisms of plants to minimize production of reactive oxygen species (ROS) at the chloroplast and to mitigate oxidative damage. Moreover, results shown here indicate that whereas production of ROS is an essential component of the hypersensitive defense response, the excessive accumulation of these toxic compounds impairs cell death containment and counteracts the effectiveness of the plant defenses to restrict pathogen infection.
Identification of Arabidopsis thaliana transformants without selection reveals a high occurrence of silenced T‐DNA integrations
Authors: Francis Kirk E. , Spiker Steven
Published: November 2008

Several recent investigations of T‐DNA integration sites in Arabidopsis thaliana have reported ‘cold spots’ of integration, especially near centromeric regions. These observations have contributed to the ongoing debate over whether T‐DNA integration is random or occurs preferentially in transcriptionally active regions. When transgenic plants are identified by selecting or screening for transgenic activity, transformants with integrations into genomic regions that suppress transcription, such as heterochromatin, may not be identified. This phenomenon, which we call selection bias, may explain the perceived non‐random distribution of T‐DNA integration in previous studies. In order to investigate this possibility, we have characterized the sites of T‐DNA integration in the genomes of transgenic plants identified by pooled polymerase chain reaction (PCR), a procedure that does not require expression of the transgene, and is therefore free of selection bias. Over 100 transgenic Arabidopsis plants were identified by PCR and compared with kanamycin‐selected transformants from the same T1 seed pool. A higher perceived transformation efficiency and a higher frequency of transgene silencing were observed in the PCR‐identified lines. Together, the data suggest approximately 30% of transformation events may result in non‐expressing transgenes that would preclude identification by selection. Genomic integration sites in PCR‐identified lines were compared with those in existing T‐DNA integration databases. In PCR‐identified lines with silenced transgenes, the integration sites mapped to regions significantly underrepresented by T‐DNA integrations in studies where transformants were identified by selection. The data presented here suggest that selection bias can account for at least some of the observed non‐random integration of T‐DNA into the Arabidopsis genome.
Hepatocellular carcinoma in a 10‐month‐old biliary atresia child
Authors: Iida Taku , Zendejas Ivan R. , Kayler Liise K. , Magliocca Joseph F. , Kim Robin D. , Hemming Alan W. , Gonzalez‐Peralta Regino P. , Fujita Shiro
Published: November 2008

Abstract:  We present a case of a 10‐month‐old boy with BA who developed HCC and was treated with liver transplantation. A four‐month‐old boy was referred to our institution because of persistent jaundice, hepatomegaly, and coagulopathy. He had been treated for the diagnosis of neonatal hepatitis at an outside hospital. He was evaluated and was accepted as a liver transplant candidate, and was subsequently transplanted with a deceased donor liver allograft at the age of 10 months. His native liver showed established cirrhosis because of BA with one focus of moderately differentiated HCC, measuring 0.7 cm in a diameter with microscopic vascular invasion in pathological study. The postoperative course was uneventful, and he is well without recurrence four months after liver transplantation. The occurrence of HCC in a child under one yr old is extremely rare, and only three cases are reported so far including our case.