bluuebunny/crossref_metadata_2025 huggingface dataset:
The combined impact ofCYP2C19andCYP2B6pharmacogenetics on cyclophosphamide bioactivation
Authors: Helsby Nuala A. , Hui Chung‐Yee , Goldthorpe Michael A. , Coller Janet K. , Soh May Ching , Gow Peter J. , De Zoysa Janak Z. , Tingle Malcolm D.
Published: August 2010

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• The prodrug cyclophosphamide requires bioactivation by liver CYP enzymes.

• Controversy exists about which CYP isoforms are important in thein vitrobioactivation of this drug.

• Recent clinical studies have highlighted a role for eitherCYP2C19orCYP2B6in the therapeutic response to cyclophosphamide in lupus patients.

• However, the role of these isoforms in the bioactivation of cyclophosphamide in lupus patients has not been previously demonstrated.

WHAT THIS STUDY ADDS

• Low bioactivation of cyclophosphamide by human liver appears to be dependent on a combination of bothCYP2C19andCYP2B6loss of function variants.

• In a preliminary study of lupus patients poor bioactivation of cyclophosphamide was also observed in those individuals who had at least one loss of function allele at eitherCYP2C19orCYP2B6.

AIMS

The role of CYP pharmacogenetics in the bioactivation of cyclophosphamide is still controversial. Recent clinical studies have suggested a role for eitherCYP2C19orCYP2B6. The aim of this study was to clarify the role of these pharmacogenes.

METHODS

We used a combinedin vitro–in vivoapproach to determine the role of these pharmacogenes in the bioactivation of the prodrug to 4‐hydroxy cyclophosphamide (4‐OHCP). Cyclophosphamide metabolism was determined in a human liver biobank (n= 14) and in patients receiving the drug for treatment of lupus nephritis (n= 16)

RESULTS

In livers of knownCYP2C19andCYP2B6genotype and protein expression we observed that there was a combined role for both CYP2C19 and CYP2B6 in the bioactivation of cyclophosphamidein vitro. The presence of at least one loss of function (LoF) allele at eitherCYP2C19orCYP2B6resulted in a significant decrease in bothVmax(P= 0.028) and CLint(P= 0.0017) compared with livers with no LoF alleles. This dual genotype relationship was also observed in a preliminary clinical study, with patients who had ≥1 LoF allele at eitherCYP2C19orCYP2B6also displaying significantly (P= 0.0316) lower bioactivation of cyclophosphamide. The mean 4‐OHCP : CP bioactivation ratio was 0.0014 (95% CI 0.0007, 0.002) compared with 0.0071 (95% CI 0.0001, 0.014) in patients with no LoF alleles at either of these genes.

CONCLUSIONS

The presence of ≥1 LoF allele(s) at eitherCYP2B6orCYP2C19appeared to result in decreased bioactivation of cyclophosphamide bothin vitroand in patients. Further clinical studies to confirm this relationship are warranted.
In vitro and in vivo oxidative metabolism and glucuronidation of anastrozole
Authors: Kamdem Landry K. , Liu Yong , Stearns Vered , Kadlubar Susan A. , Ramirez Jacqueline , Jeter Stacie , Shahverdi Karineh , Ward Bryan A. , Ogburn Evan , Ratain Mark J. , Flockhart David A. , Desta Zeruesenay
Published: August 2010

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• Anastrozole is primarily cleared by hepatic metabolism via oxidative and conjugating enzymes.

WHAT THIS STUDY ADDS

• Anastrozole is oxidized to hydroxyanastrozole mainly by CYP3A4/5 and glucuronidated to anastrozole glucuronide predominantly by UGT1A4 in vitro.

• Hydroxyanastrozole glucuronide and hydroxyanastrozole were quantified as the major metabolites of anastrozole in plasma of breast cancer patients.

• This study describes for the first time anastrozole metabolic pathways and the enzymes involved, which may serve as the scientific basis for pharmacogenetic and drug‐interaction assessments.

AIMS

Little information is available regarding the metabolic routes of anastrozole and the specific enzymes involved. We characterized anastrozole oxidative and conjugation metabolism in vitro and in vivo.

METHODS

A sensitive LC‐MS/MS method was developed to measure anastrozole and its metabolites in vitro and in vivo. Anastrozole metabolism was characterized using human liver microsomes (HLMs), expressed cytochrome P450s (CYPs) and UDP‐glucuronosyltransferases (UGTs).

RESULTS

Hydroxyanastrozole and anastrozole glucuronide were identified as the main oxidative and conjugated metabolites of anastrozole in vitro, respectively. Formation of hydroxyanastrozole from anastrozole was markedly inhibited by CYP3A selective chemical inhibitors (by >90%) and significantly correlated with CYP3A activity in a panel of HLMs (r= 0.96, P= 0.0005) and mainly catalyzed by expressed CYP3A4 and CYP3A5. The Km values obtained from HLMs were also close to those from CYP3A4 and CYP3A5. Formation of anastrozole glucuronide in a bank of HLMs was correlated strongly with imipramine N‐glucuronide, a marker of UGT1A4 (r= 0.72, P < 0.0001), while expressed UGT1A4 catalyzed its formation at the highest rate. Hydroxyanastrozole (mainly as a glucuronide) and anastrozole were quantified in plasma of breast cancer patients taking anastrozole (1 mg day−1); anastrozole glucuronide was less apparent.

CONCLUSION

Anastrozole is oxidized to hydroxyanastrozole mainly by CYP3A4 (and to some extent by CYP3A5 and CYP2C8). Once formed, this metabolite undergoes glucuronidation. Variable activity of CYP3A4 (and probably UGT1A4), possibly due to genetic polymorphisms and drug interactions, may alter anastrozole disposition and its effects in vivo.
Targeting paediatric acute lymphoblastic leukaemia: novel therapies currently in development
Authors: Lee‐Sherick Alisa B. , Linger Rachel M. A. , Gore Lia , Keating Amy K. , Graham Douglas K.
Published: August 2010

Modifications to the treatment of acute lymphoblastic leukaemia (ALL) in children have led to a dramatic increase in survival in the past 40 years. Despite this success, a significant subset of paediatric leukaemia patients either relapse or fail to ever achieve a complete remission. Additionally, some patients necessitate treatment with intensified chemotherapy regimens due to clinical or laboratory findings which identify them as high risk. These patients are unlikely to respond to further minor adjustments to the dosing or timing of administration of the same chemotherapy medications. Many novel targeted therapies for the treatment of childhood ALL provide potential mechanisms to further improve cure rates, and provide the possibility of minimizing toxicity to non‐malignant cells, given their specificity to malignant cell phenotypes. This article explores many of the potential targeted therapies in varying stages of development, from those currently in clinical trials to those still being refined in the research laboratory.
Desmoplakin and KIF20B as target antigens in patients with paroxysmal nocturnal haemoglobinuria
Authors: Alahmad Adnan , Preuss Klaus‐Dieter , Schenk Joachim , Füreder Wolfgang , Schrezenmeier Hubert , Müller‐Lantzsch Nikolaus , Schubert Jörg , Pfreundschuh Michael
Published: August 2010

At least two hypotheses have been proposed to explain the mechanism of clonal expansion of mutant cells in paroxysmal nocturnal haemoglobinuria (PNH). One hypothesis assumes an immune escape mechanism and another proposes an intrinsic second mutational event within clonal cells. We hypothesised that autoantibodies detected in PNH patients could identify antigens that might play a role in the pathophysiology of this disease and screened a human fetal liver cDNA library for serological reactivity against haematopoietic stem/progenitor cells antigens using the SEREX approach. Two antigens were identified that are constitutively expressed in CD34+ cells. Three and four of 10 PNH patients showed antibody responses against kinesin family member 20B (KIF20B) (previously termed M‐phase phosphoprotein 1, MPP1) and desmoplakin (DSP) respectively. We also found an antibody response in one of 20 healthy volunteers against desmoplakin, yet at a much lower titre than in PNH patients. No response to KIF20B or desmoplakin was detected in five patients with aplastic anaemia without a glycosyl phosphatidyl inositol ‐deficient clone. We conclude that KIF20B and desmoplakin have been shown to be the first known auto‐antigens to be recognised by the immune system of patients with PNH. The analysis of the mechanisms underlying the autoimmune response might contribute to our understanding of the clonal expansion in PNH.
Silencing of ROR1 and FMOD with siRNA results in apoptosis of CLL cells
Authors: Choudhury Aniruddha , Derkow Katja , Daneshmanesh Amir Hossein , Mikaelsson Eva , Kiaii Shahryar , Kokhaei Parviz , Österborg Anders , Mellstedt Håkan
Published: August 2010

We have previously demonstrated that ROR1 and FMOD (fibromodulin) are two genes upregulated in chronic lymphocytic leukaemia (CLL) cells compared to normal blood B cells. In this study, siRNAs were used to specifically silence ROR1 and FMOD expression in CLL cells, healthy B cells and human fibroblast cell lines. siRNA treatment induced a specific reduction (75–95%) in FMOD and ROR1 mRNA. Western blot analysis with specific antibodies for FMOD and ROR1 demonstrated that the proteins were significantly downregulated 48 h after siRNA treatment. Silencing of FMOD and ROR1 resulted in statistically significant (P ≤ 0·05–0·001) apoptosis of CLL cells but not of B cells from normal donors. Human fibroblast cell lines treated with FMOD and ROR1 siRNA did not undergo apoptosis. This is the first report demonstrating that ROR1 and FMOD may be involved in the survival of CLL cells. ROR1 in particular is further explored as potential target for therapy in CLL.
The relationship between liver disease stage and liver fibrosis: a tangled web
Authors: Germani Giacomo , Burroughs Andrew K , Dhillon Amar P
Published: August 2010

Germani G, Burroughs A K & Dhillon A P  (2010) Histopathology57, 773–784

The relationship between liver disease stage and liver fibrosis: a tangled web

The structural consequences of chronic liver disease are described as a series of liver disease ‘stages’ with scarring and architectural change that eventually destroys and replaces the normal lobular structure of the liver. Fibrosis (‘excess collagen’) and stage have been confused in histological staging systems. Fibrosis is part of increasing liver disease stage, but fibrosis and stage are different. Staging liver disease is important in routine histopathological assessment. Measurement of liver fibrosis is another process. The collagenous proportion of a liver biopsy [collagen proportionate area (CPA)] correlates with hepatic venous pressure gradient (HVPG), which is of recognized prognostic value. CPA at 1 year post‐transplantation in hepatitis C virus‐infected patients predicts subsequent clinical decompensation. CPA in cirrhotic patients predicts decompensation more accurately than staging or HVPG. The ‘cirrhosis’ stage category has poor prognostic power, and CPA effectively substages cirrhosis. CPA improves the description of liver disease stage. Proper validation of antifibrotic treatments and ‘non‐invasive markers of liver fibrosis’ requires measurement of liver fibrosis (and not liver biopsy stage scores). It is unacceptable for the words ‘fibrosis’ and ‘score’ to remain next to each other. There are benefits to properly understanding liver fibrosis and liver disease stage and properly assessing each of them.
Methylation profiles of hereditary and sporadic ovarian cancer
Authors: Bol Guus M , Suijkerbuijk Karijn P M , Bart Joost , Vooijs Marc , Van Der Wall Elsken , Van Diest Paul J
Published: August 2010

Bol G M, Suijkerbuijk K P M, Bart J, Vooijs M, van der Wall E & van Diest P J (2010) Histopathology 57, 363–370 Methylation profiles of hereditary and sporadic ovarian cancer

Aims:  Tumour suppressor gene silencing through promoter hypermethylation plays an important role in oncogenesis. Carcinogenesis of hereditary cancers usually differs from that of their sporadic counterparts, but methylation has hardly been studied in hereditary ovarian cancer. The aim of this study was to investigate promoter methylation of a set of common tumour suppressor genes in BRCA1‐related ovarian cancer in comparison with sporadic ovarian cancer.

Methods and results:  Methylation‐specific multiplex ligation‐dependent probe amplification was used to assess the extent of promoter methylation of 24 different tumour suppressor genes in BRCA1‐associated (n = 25) and matched sporadic ovarian tumours (n = 50). A cumulative methylation index (CMI) was calculated and differences between individual genes were analysed. There was no significant difference in cumulative methylation between BRCA1‐associated and sporadic ovarian carcinomas (median CMI 108; CMI 110; P = 0.86). Also, methylation patterns of individual genes did not show distinct differences after correction for multiple comparisons. CDH13, GSTP1 and RASSF1 were frequently methylated in both sporadic and hereditary ovarian cancers. BRCA1 methylation occurred in 14% of sporadic tumours, but was not detected in BRCA1‐associated tumours.

Conclusions:  CDH13, GSTP1 and RASSF1 are frequently methylated in both sporadic and BRCA1‐associated ovarian cancers. Interestingly, methylation of BRCA1, while frequent in sporadic ovarian cancer, never occurred in the hereditary group. BRCA1‐associated ovarian cancers mimic their sporadic counterparts in extent and pattern of promoter methylation of several common tumour suppressor genes. This finding could have implications for future chemotherapy regimens based on epigenetic changes.
Homogeneous EGFR amplification defines a subset of aggressive Barrett’s adenocarcinomas with poor prognosis
Authors: Marx Andreas H , Zielinski Margarete , Kowitz Charlotte‐Marie , Dancau Ana‐Maria , Thieltges Sabrina , Simon Ronald , Choschzick Matthias , Yekebas Emre , Kaifi Jussuf T , Mirlacher Martina , Atanackovic Djordje , Brümmendorf Tim H , Fiedler Walter , Bokemeyer Carsten , Izbicki Jakob R , Sauter Guido
Published: August 2010

Marx A H, Zielinski M, Kowitz C‐M, Dancau A‐M, Thieltges S, Simon R, Choschzick M, Yekebas E, Kaifi J T, Mirlacher M, Atanackovic D, Brümmendorf T H, Fiedler W, Bokemeyer C, Izbicki J R & Sauter G (2010) Histopathology 57, 418–426 Homogeneous EGFR amplification defines a subset of aggressive Barrett’s adenocarcinomas with poor prognosis

Aims:  The epidermal growth factor receptor (EGFR) is a tyrosine kinase (TK) involved in the tumour progression of many cancer types and may serve as an important therapeutic target (erlotinib, cetuximab). Heterogeneity of EGFR amplification and expression could represent a major drawback for anti‐EGFR therapy. The aim of this study was performed to determine the potential impact of tumour heterogeneity on anti‐EGFR therapy in Barrett’s adenocarcinoma (BAC).

Methods and results:  Tissue microarray (TMA) sections of 112 BAC and 45 lymph node metastases were analysed for EGFR amplification and expression using fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC). A subset of 20 samples was also sequenced for EGFR exons 18–21 and Kirsten rat sarcoma viral oncogene homologue (KRAS) exons 2–3 mutations. EGFR amplification was seen in seven (6.25%) of 112 interpretable BAC and typically high‐level with more than 10–20 EGFR copies per tumour cell (EGFR/centromere 7 ratio >3). EGFR amplification was associated with high pT, pN and poor prognosis (P = 0.0004). Identical EGFR amplification status was found in 29 primary tumours and 29 matched lymph node metastases. Moreover, FISH analysis of three to 16 large sections from all amplified BAC and corresponding lymph node metastases did not reveal any heterogeneity of EGFR amplification. No EGFR mutation but one KRAS mutation was found.

Conclusion:  The high level and homogeneity of EGFR amplification in primary tumours and metastases suggests the potential therapeutic utility of anti‐EGFR drugs in BAC.
Next‐generation small RNA sequencing for microRNAs profiling in the honey bee Apis mellifera
Authors: Chen X. , Yu X. , Cai Y. , Zheng H. , Yu D. , Liu G. , Zhou Q. , Hu S. , Hu F.
Published: August 2010

MicroRNAs (miRNAs) are key regulators in various physiological and pathological processes via post‐transcriptional regulation of gene expression. The honey bee (Apis mellifera) is a key model for highly social species, and its complex social behaviour can be interpreted theoretically as changes in gene regulation, in which miRNAs are thought to be involved. We used the SOLiD sequencing system to identify the repertoire of miRNAs in the honey bee by sequencing a mixed small RNA library from different developmental stages. We obtained a total of 36 796 459 raw sequences; of which 5 491 100 short sequences were fragments of mRNA and other noncoding RNAs (ncRNA), and 1 759 346 reads mapped to the known miRNAs. We predicted 267 novel honey bee miRNAs representing 380 182 short reads, including eight miRNAs of other insects in 14 107 583 genome‐mapped sequences. We verified 50 of them using stem‐loop reverse‐transcription PCR (RT‐PCR), in which 35 yielded PCR products. Cross‐species analyses showed 81 novel miRNAs with homologues in other insects, suggesting that they were authentic miRNAs and have similar functions. The results of this study provide a basis for studies of the miRNA‐modulating networks in development and some intriguing phenomena such as caste differentiation in A. mellifera.
Effect of bioaggregate on differentiation of human periodontal ligament fibroblasts
Authors: Yan P. , Yuan Z. , Jiang H. , Peng B. , Bian Z.
Published: August 2010

Yan P, Yuan Z, Jiang H, Peng B, Bian Z. Effect of bioaggregate on differentiation of human periodontal ligament fibroblasts. International Endodontic Journal.

Aim  To investigate the cytotoxicity of bioaggregate (BA; Innovative Bioceramix, Vancouver, BC, Canada) to human periodontal ligament (PDL) fibroblasts and its effect on differentiation of human PDL fibroblasts and to compare its performance to that of mineral trioxide aggregate (MTA; Dentsply, Tulsa, OK, USA).

Methodology  Cytotoxicity was assessed by 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay on days 1, 2, and 3 after incubation with BA or MTA. The influence of BA and MTA on differentiation of human PDL fibroblasts on days 3, 5, and 7 was evaluated by gene expression of alkaline phosphatase (ALP) and collagen type I (COLI) via quantitative real‐time polymerase chain reaction (PCR). The data were analysed by one‐way analysis of variance followed by Tukey’s test.

Results  Cell numbers in the BA group were similar to that of the control group throughout the culture period, whereas MTA suppressed the proliferation of fibroblasts. ALP expression was significantly increased in the BA group on day 7, whilst it was enhanced by MTA on day 3. Gene expression of COLI was induced by both BA and MTA compared to the control group.

Conclusions  Bioaggregate was nontoxic to human PDL fibroblasts and appeared to induce the differentiation of human PDL fibroblasts.
Microcalcifications in atherosclerotic lesion of apolipoprotein E‐deficient mouse
Authors: Debernardi Nicola , Roijers Ruben B. , Krams Rob , De Crom Rini , Mutsaers Peter H.A. , Van Der Vusse Ger J.
Published: August 2010

Evidence is accumulating that calcium‐rich microdeposits in the vascular wall might play a crucial role in the onset and progression of atherosclerosis. Here we investigated an atherosclerotic lesion of the carotid artery in an established murine model, i.e. the apolipoprotein E‐deficient (APOE−/−) mouse to identify (i) the presence of microcalcifications, if any, (ii) the elemental composition of microcalcifications with special reference to calcium/phosphorus mass ratio and (iii) co‐localization of increased concentrations of iron and zinc with microcalcifications. Atherosclerosis was induced by a flow‐divider placed around the carotid artery resulting in low and high shear‐stress regions. Element composition was assessed with a proton microprobe. Microcalcifications, predominantly present in the thickened intima of the low shear‐stress region, were surrounded by areas with normal calcium levels, indicating that calcium‐precipitation is a local event. The diameter of intimal microcalcifications varied from 6 to 70 μm. Calcium/phosphorus ratios of microcalcifications varied from 0.3 to 4.8, mainly corresponding to the ratio of amorphous calcium‐phosphate. Increased iron and zinc concentrations commonly co‐localized with microcalcifications. Our findings indicate that the atherosclerotic process in the murine carotid artery is associated with locally accumulated calcium, iron and zinc. The calcium‐rich deposits resemble amorphous calcium phosphate rather than pure hydroxyapatite. We propose that the APOE−/− mouse, in which atherosclerosis was evoked by a flow‐divider, offers a useful model to investigate the pathophysiological significance of accumulation of elements such as calcium, iron and zinc.
Erythropoietin is involved in angiogenesis in human primary melanoma
Authors: Ribatti Domenico , Nico Beatrice , Perra Maria Teresa , Longo Vito , Maxia Cristina , Annese Tiziana , Piras Franca , Murtas Daniela , Sirigu Paola
Published: August 2010

In this study, the extent of angiogenesis, evaluated as microvascular volume density, immunoreactivity of tumour cells to erythropoietin (Epo) and of endothelial cells to Epo receptor (EpoR) have been correlated in human primary melanoma specimens. Results showed that Epo/EpoR expression correlate with angiogenesis and tumour thickness. These findings suggest that Epo is secreted by tumour cells and it affects vascular endothelial cells via its receptor and promotes angiogenesis in a paracrine manner, playing an important role in melanoma angiogenesis.
Strong facilitation in mild environments: the stress gradient hypothesis revisited
Authors: Holmgren Milena , Scheffer Marten
Published: August 2010

1. The idea that the role of facilitative interactions increases as environmental conditions become more stressful has become a ruling paradigm in ecology. Here, we review three reasons why positive interactions may actually be more prominent than generally thought under moderately stressful rather than under extreme conditions.

2. First, there is evidence that in some communities the net effect of amelioration of shortage of a limiting resource, such as water under the canopy of nurse plants, may be beneficial under moderate conditions whereas it can be overruled by increased competition for the same resource in very harsh environments.

3. Secondly, we show that even in situations where the relative role of facilitation increases monotonically with stress, the absolute effect should as a rule be largest at intermediately stressful conditions. This is because under the harshest conditions, facilitative amelioration of conditions is insufficient to allow growth altogether. Therefore, while facilitation will expand the range of conditions where an organism may occur, the largest absolute effects on biomass will always occur under less stressful conditions.

4. A third reason why facilitation may be more important under moderate conditions than previously thought is that in any ecosystem, the suite of organisms is adapted to local conditions. This implies that even under conditions that appear benign, facilitation may play an unexpectedly large role as organisms are simply more sensitive than those found under harsher overall conditions.

5. Synthesis. We argue that while facilitation will extend the range of conditions where an organism can occur, it should also boost performance of the species well into the more moderate range of conditions. Broadening our search image for facilitative effects towards milder environments will reveal wider than expected prevalence of positive interactions and their effects on stability and diversity in nature.
Environment‐enhanced self‐fertilization: implications for niche shifts in adjacent populations
Authors: Levin Donald A.
Published: August 2010

1. Pollen‐mediated gene flow from ecological core populations may hamper the ability of populations to invade new, closely adjacent habitats. The potential for the invasion of a new habitat is enhanced when selection against immigrants is augmented by the elevation of self‐fertilization. This shift imparts a measure of reproductive isolation to the invader.

2. Whereas genetically mediated changes in selfing often accompany niche shifts in plants, the elevation of self‐fertilization by plastic responses to novel, stressful, environments has not been explored. Here I will show that invaders of novel habitats may have elevated levels of self‐fertility mediated by alterations in floral form and self‐compatibility.

3. Heightened selfing facilitates the evolution of local adaptation by increasing the additive genetic variance for tolerance to the new habitat. Heightened selfing also facilitates local adaptation by increasing reproductive assurance.

4. Elevated levels of selfing based on developmental responses are immediate, as opposed to that obtained by multigenerational selection.

5. Synthesis. The likelihood of a niche shift will be substantially enhanced if the invader is protected from the swamping effect of gene flow from a nearby core population. This protection may be afforded by an environment‐induced elevation of self‐fertility. The likelihood of a niche shift also is enhanced by an increase in reproductive assurance which may be vital to the survival of an invading population whose reproductive success is density‐dependent.
Effects of population size on plant reproduction and pollinator abundance in a specialized pollination system
Authors: Klank Charlotte , Pluess Andrea R. , Ghazoul Jaboury
Published: August 2010

1. Many plant species are currently experiencing negative consequences of habitat fragmentation as a result of reductions in population size and disruptions in pollination services. Plants in specialized pollination systems might be especially vulnerable to changes in plant population size and density resulting from land‐use changes.

2. Representing such a system, we chose the globeflowerTrollius europaeusL. with its pollinating flyChiastocheta, which also acts as a seed predator at its larval stage, to investigate the effects of small plant population size on reproductive success and pollinator abundance.

3. Reproductive output ofT. europaeusdeclined with increasing plant population size, whileChiastochetaabundance withinT. europaeusflowers was independent of plant population size. However, at the local level,Chiastochetanumbers within flowers were inversely correlated to localT. europaeusflower density. We further found that increasing floral densities increased plant reproductive success at the population level.

4. Chiastochetaabundance was the main driver of reproductive output ofT. europaeusthrough its dual role as an obligate pollinator and seed predator: at least someChiastochetaflies were needed to secure pollination, but a continued increase inChiastochetaflies within a flower incurred seed predation costs that greatly reduced reproductive success. Thus, high local flower density contributed positively to per capita reproductive output by dilutingChiastochetaabundance within flowers independently of overall population size.

5. Synthesis. Our findings highlight that plant population size is not always the main determinant of reproductive success for populations, but that other factors such as plant density and the specific ecology of a pollinator and its interplay with other population parameters can be more important in determining the fate of a population. Furthermore, the effects of plant population size and floral density on pollinator visitation inT. europaeusvary across scales, with implications for plant fitness. It is therefore important not to focus solely on pure plant population size in determining population viability. Thus, from a conservation perspective, even small and isolatedT. europaeuspopulations may be viable and resistant to pollination‐associated vulnerabilities depending on plant density at local (subpopulation) scales.
Conventional and advanced time series estimation: application to the Australian and New Zealand Intensive Care Society (ANZICS) adult patient database, 1993–2006
Authors: Moran John L. , Solomon Patricia J. ,   for the Adult Database Management Committee (ADMC) of the Australian and New Zealand Intensive Care Society (ANZICS)
Published: August 2010

Rationale Time series analysis has seen limited application in the biomedical Literature. The utility of conventional and advanced time series estimators was explored for intensive care unit (ICU) outcome series.

Methods Monthly mean time series, 1993–2006, for hospital mortality, severity‐of‐illness score (APACHE III), ventilation fraction and patient type (medical and surgical), were generated from the Australia and New Zealand Intensive Care Society adult patient database. Analyses encompassed geographical seasonal mortality patterns, series structural time changes, mortality series volatility using autoregressive moving average and Generalized Autoregressive Conditional Heteroscedasticity models in which predicted variances are updated adaptively, and bivariate and multivariate (vector error correction models) cointegrating relationships between series.

Results The mortality series exhibited marked seasonality, declining mortality trend and substantial autocorrelation beyond 24 lags. Mortality increased in winter months (July–August); the medical series featured annual cycling, whereas the surgical demonstrated long and short (3–4 months) cycling. Series structural breaks were apparent in January 1995 and December 2002. The covariance stationary first‐differenced mortality series was consistent with a seasonal autoregressive moving average process; the observed conditional‐variance volatility (1993–1995) and residual Autoregressive Conditional Heteroscedasticity effects entailed a Generalized Autoregressive Conditional Heteroscedasticity model, preferred by information criterion and mean model forecast performance. Bivariate cointegration, indicating long‐term equilibrium relationships, was established between mortality and severity‐of‐illness scores at the database level and for categories of ICUs. Multivariate cointegration was demonstrated for {log APACHE III score, log ICU length of stay, ICU mortality and ventilation fraction}.

Conclusions A system approach to understanding series time‐dependence may be established using conventional and advanced econometric time series estimators.
The use of statistical process control (risk‐adjusted CUSUM, risk‐adjusted RSPRT and CRAM with prediction limits) for monitoring the outcomes of out‐of‐hospital cardiac arrest patients rescued by the EMS system
Authors: Chen Tsung‐Tai , Chung Kuo‐Piao , Hu Fu‐Chang , Fan Chieh‐Min , Yang Ming‐Chin
Published: August 2010

Objective  Based on previous experience from surgical surveillance, risk‐adjusted cumulative sum (CUSUM)‐type charts were applied to monitor out‐of‐hospital cardiac arrest (OHCA) patient mortality.

Materials and methods  Data from 2356 OHCA patients were collected by the Taipei County Fire Bureau from June 2006 to November 2007. Logistic regression analysis was applied to create a risk‐adjusted model. Next, a risk‐adjusted CUSUM chart, a risk‐adjusted resetting sequential probability ratio test chart and a cumulative risk‐adjusted mortality with prediction limits chart were used to detect excess deaths of the OHCA patients rescued by the emergency medical service (EMS) system.

Results  The overall mortality rate, defined as having no return of spontaneous circulation, was 79.3%. These three charts signalled an increase in the death rate at similar sites, and also suggested a small process shift.

Conclusion  A visual approach to EMS systems monitoring that combines the risk‐adjusted cumulative sum, Risk‐adjusted resetting sequential probability ratio test and cumulative risk‐adjusted mortality with prediction limits charts was established. It was found that this approach can be effectively used by the EMS community to monitor OHCA outcomes in real time.
The role of autophagy in anticancer therapy: promises and uncertainties
Authors: Tschan M. P. , Simon H.‐U.
Published: August 2010

Abstract.  Tschan MP, Simon H‐U. (Department of Medical Oncology, Inselspital, Bern University Hospital; and Institute of Pharmacology, University of Bern; Bern, Switzerland) The role of autophagy in anticancer therapy: promises and uncertainties (Symposium). J Intern Med 2010; 268: 410–418.

Autophagy (literally self‐eating) is a catabolic mechanism involved in the recycling and turnover of cytoplasmic constituents. Although often referred to as type II programmed cell death, autophagy is primarily a survival rather than a cell death mechanism in response to different stress stimuli. Autophagy is a process in which part of the cytoplasm or entire organelles are sequestered into double‐membrane vesicles, called autophagosomes, which ultimately fuse with lysosomes to degrade their contents. Studies show that autophagy is associated with a number of pathological conditions, including cancer, infectious diseases, myopathies and neurodegenerative disorders. With respect to cancer, it has been suggested that the early stages of tumourigenesis are associated with downregulation of autophagy‐related (ATG) genes. Indeed, several ATG genes display tumour suppressor function, including Beclin1, which is frequently hemizygously deleted in breast cancer cells. Conversely, in advanced stages of tumourigenesis or during anticancer therapy, autophagy may promote survival of tumour cells in adverse environmental conditions. Therefore, a thorough understanding of autophagy in different cancer types and stages is a prerequisite to determine an autophagy‐activating or autophagy‐inhibiting treatment strategy.
Effect of Short‐and Long‐Term Gonadectomy on Neuroactive Steroid Levels in the Central and Peripheral Nervous System of Male and Female Rats
Authors: Caruso D. , Pesaresi M. , Maschi O. , Giatti S. , Garcia‐Segura L. M. , Melcangi R. C.
Published: August 2010

Significant levels of neuroactive steroids are still detected in the nervous system of rodents after the removal of peripheral steroidogenic glands. However, the influence of the plasma levels of gonadal steroids on the levels of neuroactive steroids in the nervous system has not so far been clarified in detail. Accordingly, by liquid chromatography tandem mass spectrometry, we have analysed the levels of neuroactive steroids in the sciatic nerve, in three central nervous system (CNS) regions (i.e. cerebellum, cerebral cortex and spinal cord) and in the plasma of male and female animals. The levels present in gonadally intact animals were compared with those present in short‐ and long‐term gonadectomised animals. We observed that: (i) changes in neuroactive steroid levels in the nervous system after gonadectomy do not necessarily reflect the changes in plasma levels; (ii) long‐term gonadectomy induces changes in the levels of neuroactive steroids in the peripheral nervous system (PNS) and the CNS that, in some cases, are different to those induced by short‐term gonadectomy; (iii) the effect of gonadectomy on neuroactive steroid levels is different between the PNS and the CNS and within different CNS regions; and (iv) the effects of gonadectomy on neuroactive steroid levels in the nervous system show sex differences. Altogether, these observations indicate that the nervous system adapts its local levels of neuroactive steroids in response to changes in gonadal hormones with sex and regional specificity and depending on the duration of the peripheral modifications.
Peripheral Endocannabinoid System‐Mediated Actions of Rimonabant on Growth Hormone Secretion are Ghrelin‐Dependent
Authors: Al‐Massadi O. , Gabellieri E. , Trujillo M. L . , Señaris R. , Pagotto U. , Pasquali R. , Casanueva F. F. , Seoane L . M.
Published: August 2010

The somatotroph axis is a crucial pathway regulating metabolism. Despite the fact that the endocannabinoid system has been also revealed as a potent modulator of energy homeostasis, little information is available concerning a putative interaction between these two systems. The aim of the present study was to determine thein vivoeffects of the blockade of the cannabinoid receptor type 1 (CB1) over growth hormone (GH) secretion using the CB1 antagonist rimonabant. The results obtained show that the blockade of the CB1 peripheral receptor by i.p. injection of rimonabant significantly inhibited pulsatile GH secretion. Similarly, it was found that this injection significantly decreased ghrelin‐induced GH secretion without any effect on growth hormone‐releasing hormone (GHRH)‐induced GH discharge.In situhybridisation showed that the peripheral blockade of CB1 did not affect hypothalamic somatostatin mRNA levels; however, GHRH mRNA expression was significantly decreased. The blockade of the vagus nerve signal by surgical vagotomy eliminated the inhibitory action of rimonabant on GHRH mRNA and consequently on GH. On the other hand, the central CB1 blockade by i.c.v. rimonabant treatment was unable to reproduce the effect of peripheral blockade on GHRH mRNA, nor the GH response to ghrelin. In conclusion, the data reported in the present study establish, from a physiological point of view, the existence of a novel mechanism of GH regulation implicating the action of the cannabinoid receptor on the somatotroph axis.
d‐Aspartate Oxidase Localisation in Pituitary and Pineal Glands of the Female Pig
Authors: Yamamoto A. , Tanaka H. , Ishida T. , Horiike K.
Published: August 2010

Recent evidence has shown that d‐aspartate modulates hormone secretion in the vertebral neuroendocrine system. Because only d‐aspartate oxidase (DDO) can degrade d‐aspartate, we determined DDO localisation in the pituitary and pineal glands to elucidate the control mechanisms of local d‐aspartate concentration. Brain tissues and pituitary and pineal glands of the female pigs contained a similar DDO activity of 0.38–0.66 U/g protein. However, approximately ten‐fold higher concentrations of d‐aspartate (0.27–0.35 μmol/g protein) were found in both glands. To determine the distribution of immunoreactive DDO, we made a rabbit polyclonal antibody specific to porcine DDO using a recombinant porcine enzyme. DDO immunoreactivity was found in the cytoplasm of a subgroup of cells in the anterior and intermediate lobes, in a part of nerve processes and terminals in the posterior lobe, and in the cytoplasm of a small group of pinealocytes. We used dual‐label immunocytochemistry to determine which pituitary hormones colocalise with DDO, and whether DDO and d‐aspartate immunoreactivity is reciprocal. In the pituitary gland, almost all proopiomelanocortin‐positive cells colocalised DDO, whereas only growth hormone‐positive cells colocalised d‐aspartate. d‐Aspartate immunoreactivity was not detected where DDO immunoreactivity was found. The present study suggests that DDO plays important roles to prevent undesirable off‐target action of d‐aspartate by strictly controlling local d‐aspartate concentration in the pituitary and pineal glands.
Hybridization and introgression across different ploidy levels in the Neotropical orchidsEpidendrum fulgensandE. puniceoluteum(Orchidaceae)
Authors: PINHEIRO FÁBIO , De BARROS FÁBIO , PALMA‐SILVA CLARISSE , MEYER DIOGO , FAY MICHAEL F. , SUZUKI ROGÉRIO M. , LEXER CHRISTIAN , COZZOLINO SALVATORE
Published: August 2010

The hypothesis of gene flow between species with large differences in chromosome numbers has rarely been tested in the wild, mainly because species of different ploidy are commonly assumed to be reproductively isolated from each other because of instantaneous and strong postzygotic barriers. In this study, a broad‐scale survey of molecular variation was carried out between two orchid species with different ploidy levels:Epidendrum fulgens(2n = 2x = 24 chromosomes) andEpidendrum puniceoluteum(2n = 4x = 52 chromosomes). To test the strength of their reproductive barriers, we investigated the distribution of genetic variation in sympatric and allopatric populations of these two species and conducted crossing experiments. Nuclear and plastid microsatellite loci were used to genotype 463 individuals from eight populations across the geographical range of both species along the Brazilian coastal plain. All six sympatric populations analysed presented hybrid zones, indicating that hybridization betweenE. fulgensandE. puniceoluteumis a common phenomenon. Bayesian assignment analysis detected the presence of F1and F2individuals and also signs of introgression, demonstrating a high potential for interspecific gene flow. Introgression occurs preferentially fromE. fulgenstoE. puniceoluteum. Pure parental individuals of both species display strong genotype–habitat associations, indicating that environment‐dependent selection could be acting in all hybrid zones. This study suggests that hybridization and introgression are evolutionary processes playing a role in the diversification ofEpidendrumand indicates the importance of investigations of hybrid zones in understanding reproductive barriers and speciation processes in Neotropical orchid species.
Phylogeography of ninespine sticklebacks (Pungitius pungitius) in North America: glacial refugia and the origins of adaptive traits
Authors: ALDENHOVEN JACLYN T. , MILLER MATTHEW A. , CORNELI PATRICE SHOWERS , SHAPIRO MICHAEL D.
Published: August 2010

The current geographical distribution of the ninespine stickleback (Pungitius pungitius) was shaped in large part by the glaciation events of the Pleistocene epoch (2.6 Mya–10 Kya). Previous efforts to elucidate the phylogeographical history of the ninespine stickleback in North America have focused on a limited set of morphological traits, some of which are likely subject to widespread convergent evolution, thereby potentially obscuring relationships among populations. In this study, we used genetic information from both mitochondrial DNA (mtDNA) sequences and nuclear microsatellite markers to determine the phylogenetic relationships among ninespine stickleback populations. We found that ninespine sticklebacks in North America probably dispersed from at least three glacial refugia—the Mississippi, Bering, and Atlantic refugia—not two as previously thought. However, by applying a molecular clock to our mtDNA data, we found that these three groups diverged long before the most recent glacial period. Our new phylogeny serves as a critical framework for examining the evolution of derived traits in this species, including adaptive phenotypes that evolved multiple times in different lineages. In particular, we inferred that loss of the pelvic (hind fin) skeleton probably evolved independently in populations descended from each of the three putative North American refugia.
Development of the Alaska agricultural weed flora 1981–2004: a case for prevention
Authors: CONN J S , WERDIN‐PFISTERER N R , BEATTIE K L
Published: August 2010

Conn JS, Werdin‐Pfisterer NR & Beattie KL (2011). Development of the Alaska agricultural weed flora 1981–2004: a case for prevention. Weed Research 51, 63–70.

Alaska provides a unique laboratory to study the development of agricultural weed floras, since fields were first cleared in 1900–1985. Changes in weed species composition between 1981 and 2004 were studied in 80 agricultural fields near Fairbanks, Nenana, Delta Junction and Palmer, Alaska. Cover and density of all plants were measured in quadrats along a transect in each field. Environmental and management information were collected, including field age, weed control methods, crop, elevation, latitude, longitude, surrounding vegetation type and canopy shading. Detrended correspondence analysis was used to ordinate fields based on weed vegetation. Spearman correlations and graphical overlays were used to examine relationships between environmental and management variables and ordination axes. We found seven weed species that were new to Alaskan agriculture since 1981. Crop, canopy shading, elevation, latitude and longitude were important weed flora determinants. Two distinct weed community/crop associations were identified: (i) vegetables + potatoes and (ii) perennial grass (hay + grass seed). Non‐native weed species colonised fields that were largely weed‐free in 1981, when a similar weed survey was made. The failure to use weed prevention programmes since 1981, resulted in 40 000 ha of new agricultural land that must be managed for non‐native weeds.
No loss, no gain: how deletions in our genome contribute to early‐onset obesity
Authors: Kumar RA
Published: August 2010

Large, rare chromosomal deletions associated with severe early‐onset obesity

Bochukova et al. (2010)

Nature 463(7281):666–670
Poor durability of lamivudine effectiveness despite stringent cessation criteria: A prospective clinical study in hepatitis B e antigen‐negative chronic hepatitis B patients
Authors: Liu Feng , Wang Lei , Li Xiao Ying , Liu You De , Wang Jing Bo , Zhang Zhao Hua , Wang Yao Zong
Published: August 2010

Background and Aims:  Lamivudine, a nucleoside analog, is commonly used for treatment of chronic hepatitis B (CHB) but its durability of effectiveness after withdrawal is still uncertain. This study was designed to assess the durability of lamivudine treatment with stringent cessation criteria in hepatitis B e antigen (HBeAg)‐negative patients and to explore potential predictive factors.

Methods:  Sixty one HBeAg‐negative CHB patients who had received lamivudine for at least 24 months and had maintained undetectable serum hepatitis B virus (HBV) DNA plus normal alanine aminotransferase for ≥ 18 months before withdrawal were included. They were followed up monthly during the first 4 months and at 3‐month or 6‐month intervals thereafter. Relapse was defined as serum HBV DNA ≥ 104 copies/mL.

Results:  Thirty one of 61 patients relapsed during follow‐up, over 90% occurred within 18 months after lamivudine withdrawal. Cumulative relapse rates at months 6, 12, 24, 36, 48 and 60 were 26.2%, 43.6%, 49.7%, 52.1%, 56.1% and 56.1%, respectively. Cox regression revealed that age was the only predictive factor for relapse, with lower relapse rates found in younger patients. Hepatitis B surface antigen (HBsAg) turned negative in eight patients, and none of them relapsed during follow‐up.

Conclusion:  Effectiveness of lamivudine treatment is not durable in HBeAg‐negative CHB patients even when stringent cessation criteria are adopted, with the exception of patients aged ≤ 20 years. The ideal end point of lamivudine treatment is clearance of serum HBsAg.