bluuebunny/crossref_metadata_2025 huggingface dataset:
Further evidence for an association between the gamma‐aminobutyric acid receptor A, subunit 4 genes on chromosome 4 and Fagerström Test for Nicotine Dependence
Authors: Agrawal Arpana , Pergadia Michele L. , Balasubramanian Sumitra , Saccone Scott F. , Hinrichs Anthony L. , Saccone Nancy L. , Breslau Naomi , Johnson Eric O. , Hatsukami Dorothy , Martin Nicholas G. , Montgomery Grant W. , Goate Alison M. , Rice John P. , Bierut Laura J. , Madden Pamela A.F.
Published: February 2009

Aims  A previous association analysis identified polymorphisms in gamma‐aminobutyric acid receptor A, subunit 4 (GABRA4) and GABRA2 to be associated with nicotine dependence, as assessed by a score of 4 or more on the Fagerström Test for Nicotine Dependence (FTND). In the present report, we extend the previous study by expanding our genotyping efforts significantly for these two genes.

Design  In 1049 cases (FTND of 4 or more) and 872 controls (smokers with FTND of 0) from the United States and Australia, we examine the association between 23 GABRA4 and 39 GABRA2 recently genotyped single nucleotide polymorphisms (SNPs) and nicotine dependence using logistic regression‐based association analyses using the genomic analysis package PLINK.

Results  Two and 18 additional SNPs in GABRA4 and GABRA2, respectively, were associated with nicotine dependence. The SNPs identified in GABRA4 (P‐value = 0.002) were restricted to introns 1 and 2, exon 1 and the 5′ end of the gene, while those in GABRA2 localized to the 3′ end of the gene and spanned introns 9–3, and were in moderate to high linkage disequilibrium (as measured by r2) with each other and with previously studied polymorphisms.

Conclusion  Our findings demonstrate consistently the role of GABRA4 and GABRA2 in nicotine dependence. However, further research is needed to identify the biological influence of these intronic variations and to isolate functionally relevant polymorphisms neighboring them.
Modeling the genetic and environmental association between peer group deviance and cannabis use in male twins
Authors: Gillespie Nathan A. , Neale Michael C. , Jacobson Kristen , Kendler Kenneth S.
Published: February 2009

Background  Peer group deviance (PGD) is linked strongly to liability to drug use, including cannabis. Our aim was to model the genetic and environmental association, including direction of causation, between PGD and cannabis use (CU).

Method  Results were based on 1736 to 1765 adult males from the Mid‐Atlantic Twin Registry with complete CU and PGD data measured retrospectively at three time‐intervals between 15 and 25 years using a life‐history calendar.

Results  At all ages, multivariate modeling showed that familial aggregation in PGD was explained by a combination of additive genetic and shared environmental effects. Moreover, the significant PGD–CU association was best explained by a CU→PGD causal model in which large portions of the additive genetic (50–78%) and shared environmental variance (25–73%) in PGD were explained by CU.

Conclusions  Until recently PGD was assumed to be an environmental, upstream risk factor for CU. Our data are not consistent with this hypothesis. Rather, they suggest that the liability to affiliate with deviant peers is explained more clearly by a combination of genetic and environmental factors that are indexed by CU which sits as a ‘risk indicator’ in the causal pathway between genetic and environmental risks and the expression of PGD. This is consistent with a process of social selection by which the genetic and environmental risks in CU largely drive the propensity to affiliate with deviant peers.
Impact of US and Canadian precursor regulation on methamphetamine purity in the United States
Authors: Cunningham James K. , Liu Lon‐Mu , Callaghan Russell
Published: February 2009

Aims  Reducing drug purity is a major, but largely unstudied, goal of drug suppression. This study examines whether US methamphetamine purity was impacted by the suppression policy of US and Canadian precursor chemical regulation.

Design  Autoregressive integrated moving average (ARIMA)‐intervention time–series analysis.

Setting  Continental United States and Hawaii (1985–May 2005).

Interventions  US federal regulations targeting precursors, ephedrine and pseudoephedrine, in forms used by large‐scale producers were implemented in November 1989, August 1995 and October 1997. US regulations targeting precursors in forms used by small‐scale producers (e.g. over‐the‐counter medications) were implemented in October 1996 and October 2001. Canada implemented federal precursor regulations in January 2003 and July 2003 and an essential chemical (e.g. acetone) regulation in January 2004.

Measurements  Monthly median methamphetamine purity series.

Findings  US regulations targeting large‐scale producers were associated with purity declines of 16–67 points; those targeting small‐scale producers had little or no impact. Canada's precursor regulations were associated with purity increases of 13–15 points, while its essential chemical regulation was associated with a 13‐point decrease. Hawaii's purity was consistently high, and appeared to vary little with the 1990s/2000s regulations.

Conclusions  US precursor regulations targeting large‐scale producers were associated with substantial decreases in continental US methamphetamine purity, while regulations targeting over‐the‐counter medications had little or no impact. Canada's essential chemical regulation was also associated with a decrease in continental US purity. However, Canada's precursor regulations were associated with purity increases: these regulations may have impacted primarily producers of lower‐quality methamphetamine, leaving higher‐purity methamphetamine on the market by default. Hawaii's well‐known preference for ‘ice’ (high‐purity methamphetamine) may have helped to constrain purity there to a high, attenuated range, possibly limiting its sensitivity to precursor regulation.
Gastrointestinal parasites of the grasscutter (Thryonomys swinderianus, Temminck 1827) on the Accra Plains of Ghana
Authors: Kankam Theodora , Adu Emmanuel K. , Awumbila Bawa
Published: February 2009

The gastrointestinal parasites of the grasscutter were investigated between 1996 and 2006 using 180 grasscutters from the Accra Plains. The aim of the study was to develop strategies for the control of the parasites under captive management. The gastrointestinal tracts of the animals were examined for the presence of both helminth and protozoan parasites with the aid of a hand lens, the direct smear and Willis Flotation techniques. In all, fourteen species of helminths were found including twelve nematodes and two cestodes. Trichuris muris was the most common nematode encountered, whilst Hepatocola hepatica was the least commonly encountered nematode. Three genera of protozoans, viz, Trichomonas spp, Giardia spp and Eimeria spp were also encountered, with Giardia spp and Trichomonas spp being reported for the first time in the grasscutter. Eimeria spp were the most prevalent protozoa. The widespread occurrence of protozoal and helminthic parasites in the grasscutter suggests that routine treatment of grasscutters with coccidiostats, anti‐flagellates and anthelminthics such as dimetridazole and albendazole may contribute significantly to improve productivity of animals and reduce the pressure on wild populations.
Clinical trial: dexlansoprazole MR, a proton pump inhibitor with dual delayed‐release technology, effectively controls symptoms and prevents relapse in patients with healed erosive oesophagitis
Authors: METZ D. C. , HOWDEN C. W. , PEREZ M. C. , LARSEN L. , O’NEIL J. , ATKINSON S. N.
Published: February 2009

Background  Dexlansoprazole MR heals all grades of erosive oesophagitis (EO).

Aim  To assess efficacy and safety of dexlansoprazole MR in maintaining healed EO and heartburn relief.

Methods  In this randomized, double‐blind trial, 445 patients with healed EO received dexlansoprazole MR 30 mg or 60 mg or placebo once daily for 6 months. This trial assessed maintenance of endoscopic healing (primary endpoint) and continued symptom relief based on daily diaries (secondary endpoints).

Results  Dexlansoprazole MR 30 mg and 60 mg were superior to placebo for maintaining healed EO (P < 0.0025; Hochberg’s). By life‐table analysis, maintenance rates were 75%, 83% and 27% for dexlansoprazole MR 30 mg, 60 mg and placebo respectively. Crude maintenance rates were 66% for both dexlansoprazole MR doses and 14% for placebo. Dexlansoprazole MR controlled heartburn (medians of 91–96% for 24‐h heartburn‐free days, 96–99% for heartburn‐free nights). The only more common adverse event occurring at a significantly higher rate in dexlansoprazole MR groups than placebo when analysed per patient‐months of exposure was upper respiratory tract infection.

Conclusions  Dexlansoprazole MR effectively maintained EO healing and symptom relief; most patients were heartburn‐free for >90% of days. Both doses were well tolerated.
Expression of regulatory genes for lymphoplasmacytic cell differentiation in Waldenstrom Macroglobulinemia
Authors: Leleu Xavier , Hunter Zachary R. , Xu Lian , Roccaro Aldo M. , Moreau Anne‐Sophie , Santos Daniel D. , Hatjiharissi Evdoxia , Bakthavachalam Vinod , Adamia Sophia , Ho Allen W. , Soumerai Jacob , Patterson Christopher J. , Manning Robert J. , Hamilton Susanna , Verselis Sigitas , Fox Edward , Carrasco Ruben , Ghobrial Irene M. , Treon Steven P.
Published: February 2009

Waldenstrom Macroglobulinemia (WM) is a B‐cell malignancy characterized by excess bone marrow (BM) lymphoplasmacytic cells (LPC). The accumulation of LPC in WM may represent a failure of B‐cells to properly differentiate into plasma cells. The present study investigated transcriptional expression of genes involved in late B‐cell differentiation, including PRDM1, PAX5, XBP1 transcripts and ERN1, in BM B‐cells from 31 patients with WM and six healthy donors. Real time reverse transcription polymerase chain reaction (RT‐PCR) determined that approximately 80% of the patients had high XBP1 spliced mRNA expression, 80% of whom had high mRNA ERN1α expression. XBP1, PRDM1 and PAX5 mRNA was present in all patients studied. Using relative quantitative RT‐PCR we isolated two groups with low and high expression of XBP1, XBP1 spliced and ERN1α. Sequence analysis showed germline polymorphisms in all genes studied. These data depict for the first time a heterogeneous expression pattern of the genes involved in late differentiation process of plasma cells in patients with WM and propose a role of XBP1‐ERN1α in WM pathogenesis.
Adaptation of soil microbial communities to temperature: comparison of fungi and bacteria in a laboratory experiment
Authors: BÁRCENAS‐MORENO GEMA , GÓMEZ‐BRANDÓN MARIA , ROUSK JOHANNES , BÅÅTH ERLAND
Published: February 2009

Temperature not only has direct effects on microbial activity, but can also affect activity indirectly by changing the temperature dependency of the community. This would result in communities performing better over time in response to increased temperatures. We have for the first time studied the effect of soil temperature (5–50 °C) on the community adaptation of both bacterial (leucine incorporation) and fungal growth (acetate‐in‐ergosterol incorporation). Growth at different temperatures was estimated after about a month using a short‐term assay to avoid confounding the effects of temperature on substrate availability. Before the experiment started, fungal and bacterial growth was optimal around 30 °C. Increasing soil temperature above this resulted in an increase in the optimum for bacterial growth, correlated to soil temperature, with parallel shifts in the total response curve. Below the optimum, soil temperature had only minor effects, although lower temperatures selected for communities growing better at the lowest temperature. Fungi were affected in the same way as bacteria, with large shifts in temperature tolerance at soil temperatures above that of optimum for growth. A simplified technique, only comparing growth at two contrasting temperatures, gave similar results as using a complete temperature curve, allowing for large scale measurements also in field situations with small differences in temperature.
Historical ozone concentrations and flavonoid levels in herbarium specimens of the Antarctic mossBryum argenteum
Authors: RYAN KEN G. , BURNE ALLAN , SEPPELT RODNEY D.
Published: February 2009

Depletion of stratospheric ozone since the mid 1970s has led to significant increases in ultraviolet B (UVB) irradiation over Antarctica. The detrimental effects of UVB on plants are many, but plants produce photoprotective flavonoids that reduce cellular damage. We used herbarium samples of the mossBryum argenteumcollected in Antarctica to compare the levels of flavone aglycones in plants collected before and after the formation of the ozone hole. The interpretation of historical data is difficult, because environmental conditions immediately before sample collection are unknown. Factors such as cloud cover can have a significant influence on UVB dose at ground level, modifying the flavonoid content of the specimen and adding considerable variability to the results. Nevertheless, our results revealed significant relationships between total flavone concentration and pooled year classes (P= 0.001). Furthermore, regression analysis showed a significant negative relationship of total flavone concentration and the level of ozone immediately before the time of collection (P= 0.016). In addition, the ratio of luteolin (anortho‐dihydroxylated flavone) to apigenin (a monohydroxylated flavone) increased significantly with several environmental parameters. These included (a) increasing modelled midday UVB radiation (P= 0.002), (b) increasing modelled midday UVB/PAR ratio (P< 0.001), and (c) decreasing ozone concentration (P< 0.001). We emphasise the utility of this ratio in interpreting the historical ozone trends rather than relying on changes in total flavone concentrations alone. These results illustrate that herbarium specimens may reveal historical levels of UVB radiation.
Region‐specific assessment of greenhouse gas mitigation with different manure management strategies in four agroecological zones
Authors: SOMMER SVEN G. , OLESEN JØRGEN E. , PETERSEN SØREN O. , WEISBJERG MARTIN R. , VALLI LAURA , RODHE LENA , BÉLINE FABRICE
Published: February 2009

Livestock farming systems are major sources of trace gases contributing to emissions of the greenhouse gases (GHG) nitrous oxide (N2O) and methane (CH4), i.e. N2O accounts for 10% and CH4 for 30% of the anthropogenic contributions to net global warming. This paper presents scenario assessments of whole‐system effects of technologies for reducing GHG emissions from livestock model farms using slurry‐based manure management. Changes in housing and storage practice, mechanical separation, and incineration of the solid fraction derived from separation were evaluated in scenarios for Sweden, Denmark, France, and Italy. The results demonstrated that changes in manure management can induce significant changes in CH4 and N2O emissions and carbon sequestration, and that the effect of introducing environmental technologies may vary significantly with livestock farming practice and interact with climatic conditions. Shortening the in‐house manure storage time reduced GHG emissions by 0–40%. The largest GHG reductions of 49 to, in one case, 82% were obtained with a combination of slurry separation and incineration, the latter process contributing to a positive GHG balance of the system by substituting fossil fuels. The amount and composition of volatile solids (VS) and nitrogen pools were main drivers in the calculations performed, and requirements to improve the assessment of VS composition and turnover during storage and in the field were identified. Nevertheless, the results clearly showed that GHG emission estimates will be unrealistic, if the assumed manure management or climatic conditions do not properly represent a given country or region. The results also showed that the mitigation potential of specific manure management strategies and technologies varied depending on current management and climatic conditions.
Three Types of Gonadotrophin‐Releasing Hormone Neurones and Steroid‐Sensitive Sexually Dimorphic Kisspeptin Neurones in Teleosts
Authors: Oka Y.
Published: February 2009

In general, the gonadotrophin‐releasing hormone (GnRH) neuronal systems of vertebrates consist of one group of hypothalamic neuroendocrine and one or two group(s) of extrahypothalamic neuromodulatory GnRH neurones. By taking advantage of the brains of dwarf gourami and GnRH‐green fluorescent protein transgenic medaka, the spontaneous electrical activities of all three different types of GnRH neurones have now been characterised. The hypophysiotrophic preoptic area‐gnrh1 neurones show irregular and episodic spontaneous electrical activities, whereas extrahypothalamic midbrain gnrh2 and terminal nerve‐gnrh3 neurones show regular pacemaker potentials. It is suggested that these spontaneous electrical activities are related to their different functions as neuroendocrine hormones (gnrh1 neurones) and neuromodulators (gnrh2 and gnrh3 neurones). On the other hand, recent evidence strongly suggests that the GnRH neurones are regulated by another class of peptidergic neurones, the kisspeptin neurones. The gene encoding kisspeptin (kiss1 gene) has been cloned, and the anatomical distribution of kiss1 mRNA expressing neurones (kiss1 neurones) has recently been studied in brains of several fish species. In medaka, two kiss1 neuronal populations in hypothalamic areas, called the nucleus ventral tuberis (NVT) and nucleus posterioris periventricularis (NPPv), have been identified. The NVT kiss1 neurones are sexually dimorphic in number (male >> female) under breeding conditions and are sensitive to ovarian oestrogens, whereas the NPPv kiss1 neurones are neither sexually dimorphic, nor sensitive to steroids. The steroid‐sensitive changes in kiss1 mRNA expression in the NVT occur physiologically, closely linked to the reproductive state. As in the mammalian counterpart, the medaka kiss1 neuronal system is suggested to be involved in the central regulation of reproductive functions. Interestingly, kiss2, another gene paralogous to kiss1, has been cloned in some fish species. The possible regulation of reproduction and other unknown functions by kiss1 and kiss2 neurones may be the focus of future studies.
Results of a prospective randomized controlled trial of posterior ZrSiO4‐ceramic crowns
Authors: ENCKE B. S. , HEYDECKE G. , WOLKEWITZ M. , STRUB J. R.
Published: February 2009

Summary  The aim of this randomized controlled clinical trial was to evaluate over a 5‐year period, the clinical outcome of shrinkage‐free ZrSiO4‐ceramic (KaVo Everest HPC®) full coverage crowns on posterior teeth in comparison with conventional gold crowns that served as the control. This study reports results of a 5‐year study protocol up to 24 months. Patients totalling 224 were randomly divided into two treatment groups. Of these, 123 patients were restored with 123 Everest HPC® crowns, fabricated by the Kavo Everest computer‐aided manufacturing (CAM) procedure and 101 patients received 101 gold crowns, but two were excluded from analysis. All crowns were conventionally cemented with glass–ionomer cement. After an observation period of 6, 12 and 24 months, the prospective survival rates (Kaplan–Meier) for the KaVo Everest HPC® crowns were 97·9%, 95·1% and 89·8% and for the gold crowns 100%, 94·8% and 92·7%, respectively. There were no significant differences between the two groups (P = 0·2). The 1‐year failure rates were 4·9% for the KaVo Everest HPC® crowns and 5·2% for the gold crowns. The 1‐year cumulative risks for loss of vitality, secondary caries, fractures, loss of crown and extraction of abutment of the analyzed abutments (88) were 8·9%, 0%, 0%, 1·1% and 1·1%, respectively, for the gold crowns and 2·8%, 0%, 4·7%, 0% and 0·9%, respectively, for the ceramic crowns (107 analyzed abutments). No perfect marginal fit was shown by 49·5% of the evaluated ceramic crowns and 26·1% of the gold crowns. Only 1·9% of the KaVo Everest HPC® crowns had a marginal crevice. In conclusion, Everest HPC® crowns with an adequate occlusal tooth reduction of >1·5 mm are suitable for posterior restorations, but the marginal fit shows a potential for improvement.
An exploration of the social identity of mental health inpatient service users
Authors: JACKSON L. , TUDWAY J. A. , GILES D. , SMITH J.
Published: February 2009

Social identity theory proposes that a threatened social identity can impact on self‐concept and well‐being. As a low‐status minority group, mental health service users face the possibility of a threatened social identity compounding existing mental health problems. This may be further complicated by an inpatient admission where the context in which social identity is maintained has changed from a community setting. We use a qualitative approach to explore the salient social identity of mental health inpatient service users using semi‐structured interviews. Analysis showed that the salient in‐groups and out‐groups mainly centred around the inpatient context but were somewhat influenced by non‐mental health service group membership. Participants also talked of their unique characteristics as group members. How these findings relate to social identity theory and, particularly how they relate to mental health service users within changing contexts is explored.
Trends of hepatitis A hospitalizations and costs associated with the hospitalization in Spain (2000–2005)
Authors: Arteaga A. , Carrasco‐Garrido P. , De Andres A. L. , De Miguel Á. G. , Jiménez‐García R.
Published: February 2009

Summary.  The objective of this study was to analyse data related to hospitalization, comorbidities, average stays and costs associated with the hospitalization cases of hepatitis A in Spain, during the period between 2000 and 2005. A retrospective, descriptive study of the epidemiological characteristics of hepatitis A patients admitted to Spanish hospitals was performed using, as an information source, the Minimum Basic Data Set. National hospitalization rates were calculated for hepatitis A. Comorbidities, mortality, average and median stays, and mean medical costs related to hospitalization were analysed. Costs were calculated using Diagnosis‐Related Groups for the disease. The total number of hospitalized patients with hepatitis A diagnosis was 2351 subjects (rate 1.87/100 000 inhabitants). The highest rate corresponds to the group aged between 20 and 39 years (3.07/100 000 inhabitants). Some 60.1% of hospitalized hepatitis A cases were diagnosed in men. Twenty‐two deaths (0.9%) out of the total of hospitalized subjects were reported. The average hospital stay caused by hepatitis A was 6.8 days. The trends in the rate of hospitalized hepatitis A cases in Spain, from the year 2000 to 2005, were not statistically significant. Differences were found neither in the hospitalization percentage, nor in the average length of stay. An increase of cost from €836 278 in the year 2000 to €1 272 608 in the year 2005 was observed. The rate of hospitalized hepatitis A subjects in Spain has not changed over the period 2000–2005. The total cost derived from these hospitalizations has increased by 52%.
Prevalence of glucose intolerance in patients with chronic hepatitis B or C: a prospective case–control study
Authors: Mavrogiannaki A. , Karamanos B. , Manesis E. K. , Papatheodoridis G. V. , Koskinas J. , Archimandritis A. J.
Published: February 2009

Summary.  Despite several studies, the association of glucose intolerance with chronic hepatitis B (CHB) or C (CHC) virus infection remains controversial. We evaluated the prevalence of glucose intolerance by oral glucose tolerance test (OGTT) in patients with CHB or CHC in comparison with matched controls. In total, 189 consecutive outpatients with CHB or CHC and 189 subjects individually matched for age, sex and body mass index (BMI) were included. OGTT was performed in all cases, except in known diabetics, and glucose intolerance was defined as impaired glucose tolerance (IGT), OGTT‐diabetes or known diabetes. Most patients with abnormal OGTT had normal fasting glucose (IGT: 69.8%, OGTT‐diabetes: 54.5%). Compared with their own controls, CHB patients had a higher prevalence of IGT (13.6%vs 2.5%, P = 0.018) and family history of diabetes (34.6%vs 16.0%, P = 0.011), while CHC patents had higher prevalence of glucose intolerance (37.0%vs 15.7%, Ρ = 0.001), mostly because of more frequent IGT (21.3%vs 6.5%, Ρ = 0.003). After age and BMI adjustment, patients with CHC compared with those with CHB had significantly higher prevalence of glucose intolerance (37.0%vs 29.6%, P = 0.037). In conclusion, increased prevalence of glucose intolerance is documented by OGTT both in CHC and CHB patients compared with age, sex and BMI matched controls. Glucose intolerance is more frequent in CHC than CHB patients, regardless of known risk factors. An OGTT might be necessary at the baseline work‐up of CHB or CHC patients, as a normal fasting glucose value does not exclude IGT or OGTT‐diabetes.
Regulation and function of root exudates
Authors: BADRI DAYAKAR V. , VIVANCO JORGE M.
Published: February 2009

Root‐secreted chemicals mediate multi‐partite interactions in the rhizosphere, where plant roots continually respond to and alter their immediate environment. Increasing evidence suggests that root exudates initiate and modulate dialogue between roots and soil microbes. For example, root exudates serve as signals that initiate symbiosis with rhizobia and mycorrhizal fungi. In addition, root exudates maintain and support a highly specific diversity of microbes in the rhizosphere of a given particular plant species, thus suggesting a close evolutionary link. In this review, we focus mainly on compiling the information available on the regulation and mechanisms of root exudation processes, and provide some ideas related to the evolutionary role of root exudates in shaping soil microbial communities.
17. Immunoelectrophoretical Quantitation of Human Placental Lactogen Hormone (HPL)
Authors: NøRGAARD‐PEDERSEN B. , GæDE P.
Published: February 2009

Summary. A simple rocket‐immunoelectrophoretic method suitable for quantitation of human placental lactogen hormone in sera or plasma samples from pregnant women is described. The samples are used undiluted since the corcentration range is 1–8 mg/l. A close correlation was found between the present method and a commercially available radioimmunoassay. The day‐to‐day reproducibility in terms of relative standard deviation was found to be 4.6%. The mean half life of HPL in 3 patients just after delivery was found to be 16 minutes. Some examples of clinical applications are given in curve form.
A Simple and Reproducible Method for Crossed Immunoelectrofocusing
Authors: SöDERHOLM J. , SMYTH C. J. , WADSTRöM T.
Published: February 2009

Crossed immunoelectrofocusing comprises isoelectric focusing in the first dimension in combination with electrophoresis into antibody‐containing gel in the second dimension. Isoelectric focusing on a thin layer polyacrylamide gel permits better comparison of a number of different samples in the same run compared to isoelectric focusing in polyacrylamide rods. Strict control of several parameters during electrofocusing, slicing of the acrylamide gel and transfer of the strips to an agarose gel were found to be important factors in obtaining reproducible results. Attempts to mould acrylamide strips into agarose gels were abandoned when a more convenient way of achieving acrylamide‐agarose contact, referred to as the laying‐on method, was devised, which gave highly reproducible results. In this technique the acrylamide strip was turned upside down after electrofocusing and applied to the surface of an antibody‐containing agarose gel. Crossed immunoelectrofocusing was used to study different preparations of diphtheria toxin from three laboratories. All preparations contained toxin with pronounced microheterogeneity as evidenced by fused immunoprecipitation patterns. A possible explanation for the microheterogeneity of diphtheria toxin is that 'nicked' toxin may contain up to six different molecular species.
The Use of Matrix‐Bound Protein A from Staphylococcus aureus for the Isolation and Determination of Immunoglobulins
Authors: HJELM H. , SJöQUIST J.
Published: February 2009

Summary. In the present paper protein A (S. aureus) covalently bound to Sepharose 4B was used for the direct isolation of IgG from human serum. The protein A‐Sepharose, containing 4 mg protein per ml packed gel, could bind 20 mg IgG. The recovery of pure IgG eluted from the column was about 85%. Monovalent protein A fragments (molecular weights 6,000–8,000 daltons) covalently linked to Sepharose were used to study contaminants in some commercially available IgG preparations. Most of these contaminants (3.4 to 14.2%) were shown to be split products of IgG and in some preparations up to 2.2% IgA was present. A procedure was also developed for the determination of as little as 0.001% IgG in highly concentrated albumin solutions. Finally protein A‐Sepharose was used to isolate IgG3 from normal human sera and from a patient with multiple myelomatosis. Other practical applications of affinity chromatography using protein A‐Sepharose are also discussed.
The Danish Testicular Carcinoma Project (DATECA)
Authors:   THE DATECA STUDY GROUP
Published: February 2009

Since January 1st, 1976, almost all new cases of germ cell carcinoma of the testis occurring in Denmark have been treated on a joint protocol. Depending on stage and histology, patients receive the same post‐orchiectomy, radiotherapy and chemotherapy throughout the country. The treatment guide‐lines adopted since January 1st, 1976, are presented.
A Report of Two Cases of Primary Intracranial Germ Cell Tumours with Elevated Levels of Alpha‐fetoprotein and Human Chorionic Gonadotropin
Authors: NØRGAARD‐PEDERSEN B. , ALBRECHTSEN R. , ARENDS J. , LINDHOLM J. , SANDBERG E.
Published: February 2009

Two cases of primary intracranial germ cell tumours are reported. Both serum alpha‐fetoprotein (AFP) and serum human chorionic gonadotropin (hCG) were raised preoperatively, but became normal during postoperative radio‐ and chemotherapy. Immunofluorescence microscopy of tissue specimens from both tumours showed a slight staining for AFP mainly localized in the endodermal vesicles, whereas a more intensive staining for hCG could be identified in the giant cells.
Initial Attempt to Develop an AFP Gene Regulation Analysis System
Authors: BÉLANGER L. , COMMER P. , HARDY K. J. , CHIU J. F.
Published: February 2009

This is a preliminary report on methods being developed for the isolation of rat AFP messenger RNA and the synthesis of DNA complementary to this mRNA. Using Morris hepatoma 7777 as source material, AFP‐synthesizing polysomes have been isolated by indirect immunoprecipitation. The 18S Poly A + RNA (putative mRNAAFP) recovered from these polysomes by differential centrifugation and oligo‐dT cellulose chromatography, was translated in a reticulocyte cell‐free system into a peptide immunoprecipitable by anti‐AFP sera (and not by anti‐albumin sera) and co‐migrating with native AFP on polyacrylamide gel electrophoresis. DNA complementary to the mRNA was synthesized by the use of avian myeloblastosis virus reverse transcriptase. The cDNA probe is highly homogeneous, as judged by alkaline sucrose gradient sedimentation, and yields a 1/2 C,t value of 8.5 times 10‐3 with its template. Preliminary hybridization experiments indicate that sequences hybridizable to the cDNA probe constitute 3.0%, 0.5%, and 0.008% of the total poly A + polysomal RNA populations of 13 day old rat liver, Morris hepatoma 7777 and adult rat liver, respectively.
Analysis of Serum Alpha‐fetoprotein Dynamics in Early Infancy
Authors: TSUCHIDA Y. , KANEKO M. , SAITO S. , ENDO Y. , SHIRAKI K. , OHMI K.
Published: February 2009

In early infancy, the serum alpha‐fetoprotein (AFP) level remains quite high and the clinical interpretation of hyper‐alphafetoproteinemia in this age should be made in comparison with its normal ranges. We presented some illustrative cases of early infancy, whose AFP values were formerly considered abnormally high, but later found within normal ranges. Our normal ranges were obtained by statistical calculations of serum AFP values studied in normal subjects: namely infants with inguinal hernia and neonates at birth (the cord blood was used).
Effect of Purified Preparations of Mouse Alpha‐fetoprotein (AFP) on in vitro Induced Lymphocyte Blastogenesis
Authors: HASSOUX R. , POUPON M. F. , URIEL J.
Published: February 2009

Mouse amniotic fluids (MAF) were tested for their inhibitory effect on the transformation of lymphocytes by Concanavalin A (Con A), Phytohemagglutinin (PHA) or Lipopolysaccharide of Escherichia coli (LPS), and on mixed lymphocyte cultures (MLC). Eight out of the thirteen samples tested contained inhibitory activity equal or superior to 50% of the controls. Pure preparations of mouse alpha‐fetoprotein (AFP) were obtained from these or other MAF pools by immunoadsorption and/or by affinity chromatography on oestradiol‐Sepharose. None of the isolated samples of AFP, whether originating from immunosuppressive MAF pools or from pools lacking activity, exhibited a significant inhibitory effect on the in vitro transformation of lymphocytes. The suppressive properties of the MAF samples and the non suppressive activity of samples of pure AFP were not affected by the addition of oestradiol‐17β or after removal of endogenous oestrogens by exhaustive dialysis against 15% dioxan solutions.
Inhibition of Human Lymphocyte Transformation by Human α‐fetoprotein (HAFP): Studies on the Mode of HAFP Action and the Role of HAFP Polymorphism
Authors: LESTER E. P. , MILLER J. B. , YACHNIN S.
Published: February 2009

To explain the mechanism of inhibition of human lymphocyte transformation by human alpha‐fetoprotein (HAFP), we sought for, and failed to find, evidence of physical association between HAFP and phytomitogens or antihuman thymocyte antiserum. In addition, 20–40 fold increases in mitogen dose do not reverse the inhibition of lymphocyte transformation by a constant dose of HAFP. The presence of HAFP does not interfere with the attachment of 125I‐labelled phytohemagglutinin to the lymphocyte surface. Two‐dimensional crossed immunoelectrophoresis of HAFP isolated from the body fluids of hepatoma patients demonstrates three charged species of HAFP designated as HAFP‐1 (the most cathodal), HAFP‐2, and HAFP‐3 (the most anodal). The potency of HAFP isolates in inhibiting lymphocyte transformation can be positively correlated with the ratio HAFP‐3: HAFP‐1 in each preparation. Passage of HAFP isolates over CM‐cellulose allows the isolation of two HAFP fractions, one at pH 4.95, and the other at pH 5.24. The pH 4.95 CM‐cellulose isolate is enriched in the more electronegative HAFP species (HAFP‐3) and is, on the average, two times more potent than the pH 5.24 CM‐cellulose HAFP isolate. The latter, by comparison with native HAFP, is enriched in the more electropositive HAFP species (HAFP‐1). The CM‐cellulose HAFP isolates are identical in sialic acid content. We have isolated HAFP from the serum, ascitic fluid, and saline extract of tumor from a single hepatoma patient, and from an homogenate of fetal livers. When tested for their capacity to inhibit human lymphocyte transformation in vitro, tumor and fetal liver HAFP were found to be extremely potent; serum HAFP had intermediate potency, and ascitic fluid HAFP was the least potent. Analysis of these HAFP isolates by isoelectric focusing in polyacrylamide gels containing 8 M urea revealed at least 6 molecular HAFP variants. The proportion of one of these variants, termed HAFP‐3a, in each isolate was correlated with the immunosuppressive potency of the isolate. The sialic acid content of the various HAFP isolates did not vary significantly. Our data suggest that a post‐synthetic modification of HAFP occurs, which modulates its immunosuppressive potency.
Suppression of in vitro Parameters of the Cell‐mediated Immunity by Human Alpha‐1‐fetoprotein
Authors: AUER J. O.
Published: February 2009

A possible immunoregulatory role of human alpha‐1‐fetoprotein (HAFP) was investigated, since in contrast to mouse AFP such function has been denied for HAFP. HAFP enriched fractions as well as pure HAFP were obtained by means of two different procedures. After passage of HAFP containing ascites of patients with primary liver carcinoma on an anti‐HAFP immunosorbent column, the retained proteins were eluted first by a glycine‐NaOH‐buffer (HAFP 1) and second by NaSCN (HAFP II). HAFP was further purified by passage of the HAFP containing fractions on an anti‐whole human serum immunosorbent column, which resulted in semipurified HAFP I and II. Pure HAFP was obtained by recycling on the anti‐whole‐human serum immunosorbent column as well as by final Sephadex G‐100 gel filtration. At concentrations of 150–200 μg/ml, only HAFP I, semipurified as well as pure, exerted consistently a profound suppressive effect on the mixed leukocyte reaction as well as on the Concanavalin A induced lymphocyte proliferation, while it usually failed to do so in PHA stimulated lymphocyte cultures. At concentrations which were immunosuppressive, pure HAFP I showed no influence on the proportion of E‐rosetting lymphocytes. HAFP II did not exert comparable immunoregulatory effects.
Heterogeneity of Human Alpha‐fetoprotein (HAFP) as Revealed by Agarose Gel Electrophoresis and Iso‐electric Focusing in Ureaacrylamide Gels
Authors: LESTER E. P. , MILLER J. B. , YACHNIN S.
Published: February 2009

HAFP was purified from five patients with hepatoma, one with gastric cancer, and one with an embryonal cell tumor, as well as from fetal liver and a monkey tumor cell line grown in tissue culture. The pattern of microheterogeneity of purified HAFP was defined for each HAFP isolate, and was demonstrated to be present in native sera, using crossed immunoelectrophoresis in agarose gels and isoelectric focusing in polyacrylamide gels containing 8 M urea. Three, and in one case four, species were seen in agarose, and were further resolved to reveal 6 major species with isoelectric focusing which could be correlated with the agarose gel variants. We have demonstrated a relationship between the immunosuppressive potency of certain HAFP preparations and the proportion of specific HAFP isomers which they contain as shown by these techniques. We have desialylated each of our preparations and demonstrated that this does not alter immunosuppressive potency but leaves residual complex microheterogeneity. Desialylated HAFP isolates contain six major HAFP isomers by isoelectric focusing, indicating that HAFP heterogeneity is based upon multiple charge differences in the HAFP molecule, apart from sialic acid content. The nature of these charge differences remains to be determined. We postulate that these charge differences modulate the immunosuppressive potency of HAFP.
Purification of Ovine Alpha‐fetoprotein by Preparative Electrophoresis
Authors: LAI P. C. W. , HAY D. M. , LORSCHEIDER F. L.
Published: February 2009

Ovine alpha‐fetoprotein (AFP) was successfully purified from fetal lamb serum using preparative electrophoresis. Fetal serum (83 days of gestation) contained 24.8 mg total protein/ml and 1.04 mg AFP/ml. Approximately 0.74 mg AFP was obtained upon fractionation of 1 ml of serum. The AFP preparation was found to be homogeneous by analytical polyacrylamide gel electrophoresis at pH 6.0, 7.2 and 9.3, and by immunoelectrophoresis. The purified AFP was immunochemically related to human AFP when tested against chicken anti‐human AFP by double diffusion.
Clinical Evaluation of Combined Determination of AFP and LP‐X in a 4 Year Prospective Study
Authors: WEISS W. , EDER G. , HANAK H. , KROISS A. , NEUMAYR A.
Published: February 2009

In a prospective study 4,000 patients of a gastroenterologic and haematologic department were screened for AFP and LP‐X. X‐ray, endoscopic and scintigraphic examinations concerning liver and bile ducts were performed. Only histologically confirmed results were used for the evaluation of this prospective study. Applying combination of these methods enabled the differential diagnosis of localized liver diseases and increased additional information in cases of parenchymal damage of the liver up to 35%.
Antigenic Differences of CEA in Colonic Cancer, Gastric Cancer and Fetal Tissues
Authors: MORI T. , SHIMANO T. , LEE P.‐K. , FUJIMOTO N. , KOSAKI G.
Published: February 2009

CEA samples were prepared individually from liver metastases of gastric and colonic cancer, and purified with gel filtrations, block electrophoresis and Con A affinity chromatography (the latter was not performed in all cases). Anti‐CEA antisera were produced in 7 and 4 guinea pigs against CEA of gastric cancer and colonic cancer respectively. Upon extensive absorptions with PCA extracts of normal tissues (rich of NCA) and NCA‐2, their anti‐CEA activities were examined on micro‐Ouchterlony plates and by immunofluorescence techniques.

There were two types of anti‐CEA antibodies, as observed in our previous study [9], in which crude samples of CEA were utilized as immunogens: a type that showed identical reaction to CEA of adenocarcinomas of the digestive system, squamous cell carcinomas of endodermally derived organs, medullary carcinoma of the thyroid as well as tissues of fetal gastro‐intestinal tract but not to others, while the other type was proven to be specific to gastric or colonic cancer depending on cancer tissues from which the immunized CEA was extracted. In immunofluorescence study, no reaction of these organ‐specific anti‐CEA antibodies to fetal tissues was observed.

Further, CEA‐M (Con A binding fraction) and CEA‐P (Con A non‐binding fraction) separated by Con A affinity chromatography, showed identical reaction to any types of anti‐CEA antibodies, though CEA‐P was apparently less antigenic.
Amicon Filtration Technique in a Modified CEA‐Roche Test
Authors: ØRJASÆTER HARALD
Published: February 2009

Amicon MC 40 multi‐cell filtration system was evaluated for routine use, to avoid the time‐consuming and delaying dialysis in our modified CEA‐Roche test.

Four control samples were repeatedly tested by the two modifications, each with 10–12 different assays, and 100 plasma samples from cancer patients were run in parallel. The correlation between results obtained with the Amicon filtration technique and our standard method with dialysis was good over the entire range of measurement, from 1–16 μg/1. No significant differences were seen with the two methods. There was also little difference in the reproducibility. Standard deviations of ±0.3, ±0.4, ±0.8, ± 1.2 and ±0.3, ±0.5, ±0.5, ± 1.7 respectively were found in different ranges of measurement.

In our hands the conventional CEA‐Roche test used with the Amicon filtration technique was found to give too low values. Similar observations were made previously with dialysis and use of EDTA CEA‐standard. Our modification, which includes a plasma CEA‐standard, improved the long‐term reproducibility of the test.
Detection of Carcinoplacental Alkaline Phosphatase by Polyacrylamide Gradient Gel Electrophoresis
Authors: MASUZAWA M. , KAMADA T. , ABE H. , SAKOYAMA Y. , OKUBO S.
Published: February 2009

Human serum alkaline phosphatase (E.C. 3.1.3.1.) was subclassified into several isoenzymes including carcinoplacental alkaline phosphatase isoenzyme (CPALP) by electrophoretic procedures. Since CPALP has three subtypes, such as Regan, Nagao and Variant type, their properties are somewhat different among them, and have no specific property, no simple method has yet been ascertained to detect CPALP separately.

CPALP was detected as the fastest migrating isoenzyme band to the anode, using the polyacrylamide gel electrophoresis of more than 8.0% gel concentration. Thus, a polyacrylamide gradient gel electrophoresis of 16–8% gel concentration in Tris‐borate buffer system was introduced, which resulted in CPALP being found to be the fastest migrating isoenzyme band to the anode and microheterogeneities of each CPALP were also identified: CPALP (Variant) was electrophoresed rather faster to the anode, CPALP (Nagao); intermediate, and CPALP (Regan), as well as placental alkaline phosphatase isoenzyme; rather slower. In addition, each isoenzyme of CPALP and placental alkaline phosphatase further differentiated clearly into a few subbands.

Slow moving new bands appeared by Triton X‐100 treatment.
Diagnostic and Follow‐up Studies on CEA, AFP and ALP4 Isoenzyme in the Sera of Gynecological Malignancies
Authors: TOTTORI K. , SATO Y. , TAKEUCHI S.
Published: February 2009

Simultaneous determination of CEA, AFP and ALP4 was made in patients with various gynecological malignancies. CEA was positive in 65%, AFP was positive in 1.8% of 55 cases with cervical carcinoma stage 0—‐II. ALP4 was positive in one patient in these stages. In the group of cervical carcinoma stage III, stage IV and recurrence, CEA and AFP were positive in 100% and in 8.6% of 35 cases respectively. ALP4 was positive in 14.3%. High levels and/or progressively increasing CEA with positive ALP4 were found to be significant in poor prognostic patients in whom ALP4 was not always found positive. Out of 17 cases with endometrial carcinoma, CEA was positive in 12 patients. AFP and ALP4 were each positive in one case. Of 30 cases with ovarian carcinoma, CEA was positive in 70%, AFP in 23% and ALP4 in 14%. Increasing CEA and/or increasing AFP appeared to be correlated with poor prognosis. In the group of 5 vulvar carcinoma, only CEA was positive in 80% of the patients. Of 19 cases of choriocarcinoma, CEA was positive in 11%, AFP was positive in 5.3% and ALP4 was positive in 5.6%. There existed reverse‐correlation between serum CEA and AFP in some cases.
Modulation of HeLa Cell Alkaline Phosphatase Isoenzyme Expression by Prednisolone
Authors: KOTTEL R. H. , FISHMAN W. H.
Published: February 2009

Various HeLa cell lines have been examined in order to determine the effect of hormone treatment upon alkaline phosphatase isoenzyme expression. Both biochemical and immunoelectrophoretic analyses show that three cell lines demonstrating activity of the placental‐type (Regan) isoenzyme in absence of prednisolone are induced to much higher levels after growth in hormone‐containing medium for 72 hours. Prednisolone‐treatment resulted in increased enzyme activity but no change in the isoenzyme profile as determined by microzone and polyacrylamide disc gel electrophoresis. On the other hand, the majority of the activity in HeLa line D98AH2 in absence of hormone was found to be heat‐sensitive and inhibited by phenylalanine. Immunoelectrophoretic analysis indicated that amnion/intestinal isoenzyme was expressed. No appreciable change in total enzyme activity occurred in presence of hormone despite the induction of the Regan isoenzyme and the diminution of amnion/intestinal type. Thus, treatment of this cell line with hormone resulted in differential expression of the amnion/intestinal and placental isoenzyme components. These results support a regulatory function of prednisolone with respect to expression of alkaline phosphatase isoenzymes in tumor cell lines and identify a HeLa cell line, D98AH2, which elaborates the amnion/intestinal isoenzyme.
Limitation in the Specificity of the Assay of Human Chorionic Gonadotropin Using β Subunit Antiserum
Authors: ARENDS J.
Published: February 2009

In order to ensure a specific radioimmunoassay for human chorionic gonadotropin (hCG), suppliers of hCG‐β subunit antiserum advocate to ignore the low‐dose region of the standard curve, hence limiting the sensitivity of the assay.

Serum from 37 women with high LH levels in the menopausal range was assayed and the calculated hCG levels were based on the low‐dose region of the hCG standard curve, thus overriding the above‐mentioned recommendation.

With this modification of the method a significant correlation was seen between LH and hCG values.

The regression line relating LH in the serum samples to hCG was parallel with that relating pure pituitary LH to hCG in the hCG‐β assay.

These findings indicate that the hCG‐β assay has a limited specificity and that cross‐reactions from LH are bypassed by lowering the sensitivity of the assay.