bluuebunny/crossref_metadata_2025 huggingface dataset:
50 INVESTIGATION OF THE EFFECT OF BUTYROLACTONE I AND CYCLOHEXIMIDE TREATMENT DURING IN VITRO MATURATION OF SWINE OOCYTES
Authors: Marques M.G. , Gerger R.P.C. , Nascimento A.B. , Oliveira V.P. , Simoes R. , Lucio C.F. , Visintin J.A.
Published: December 2004

Butyrolactone I and cycloheximide specifically inhibit MPF activation and prevent the resumption of meiosis. The aim of this study was to investigate the kinetics of in vitro maturation of butyrolactone I- and cycloheximide-treated swine oocytes in an attempt to produce cytoplasts for nuclear transfer. Oocytes from slaughterhouse ovaries were randomly allotted to one of 3 treatments; group 1 (n = 102 – control – 22 hours of in vitro maturation in TCM199 supplemented with 3.05 mM glucose, 0.91 mM sodium pyruvate, 10% follicular fluid, 0.57 mM cysteine, 10 ng/mL EGF, 10 IU/mL eCG and 10 IU/mL of hCG and 22 hours of culture); Group 2 (n = 191 – blocking for 10 hours in 12.5 M butyrolactone I and in vitro maturation for 44 hours); and Group 3 (n = 175 – blocking for 10 hours in 5 M cyclohexemide and in vitro maturation for 44 hours). After in vitro maturation, oocytes were fixed and stained for evaluation of meiotic division. The percentage of oocytes at metaphase II (MII) in Groups 1 and 3 (75.49% and 70.29%, respectively) were higher (P < 0.05) than Group 2 (63.35%). Based on these results and in order to increase enucleation rates, we also investigated the proximity of the first polar body (PB) with the metaphase plate (MP) in Groups 1 and 3. After in vitro maturation (36, 40, and 44 hours), oocytes were gently decumulated and incubated in microdroplets (50-μL) of bisbenzimide solution (5 g/mL) to analyze the MP and PB positions. Group 1 (control) at 44 hours of maturation (47.05% – 48/102) and Group 3 at 40, and 44 hours (60.20% – 59/98 and 55.46% – 61/110, respectively) showed similar rates, that were higher (P < 0.05) than Group 1 at 36 hours and 40 hours (4% – 4/100 and 36% – 36/100, respectively) and Group 3 at 36 hours (34.58% – 37/107). In conclusion, Group 1 at 44 hours and Group 3 at 40 or 44 hours provide the best oocytes for enucleation because they showed a high number of matured oocytes with the first polar body and the metaphase plate located proximally.
This work was supported by FAPESP 02/10747-1.
67 PREMATURE CHROMOSOME CONDENSATION IS NOT ESSENTIAL FOR BOVINE SOMATIC NUCLEAR REPROGRAMMING
Authors: Sung L.Y. , Du F. , Jeong B.S. , Chang C.C. , Xu J. , Tian X.C. , Yang X.
Published: December 2004

The objective of this study was to determine the effect of donor nuclear exposure in MII oocyte cytoplasm on nuclear reprogramming events and subsequent development of cloned embryos in cattle. Somatic nuclear transfer (NT) was performed by electrofusion of the enucleated MII oocytes with cultured cumulus cells by ovum pickup. Donor cell-cytoplast pairs were fused by applying two direct current pulses at 2 kV/cm for 10 μs. Fused NT embryos were randomly divided in to Treatment A (immediate activation) and Treatment B (delayed activation, 4-h exposure in MII cytoplast before activation). In both treatment groups, the activation protocol was identical and consisted of incubation in cycloheximide (10 μg/mL) plus cytochalasin D (2.5 μg/mL) in M199 + 7.5% FBS for 1 h, followed by culture in cycloheximide (10 μg/mL) for an additional 4 h. Reconstructed embryos in both groups were subsequently cultured in CR1aa in 5% CO2, 5% O2, and 90% N2 at 39°C. Samples from both treatments were fixed at 0, 1, 2, 4, 6, 12, 18, 24, 30, 36, and 44 h after fusion. All fixed samples were double stained for tubulin and DNA, and observed with a laser-scanning confocal microscope for changes in nuclei and microtubules. The experiment was replicated three times. Cleavage rate and blastocyst rate were recorded and analyzed by Studen's t-test (SPSS 11.0, Chicago, IL). The staining revealed an absence of premature chromosome condensation (PCC) in all embryos in Treatment A. However, delayed activation (Treatment B) resulted in a high incidence of PCC, probably due to high levels of MPF in the MII cytoplasts. Chromosome condensation was observed in Treatment B at 4 h (82%, n = 17), 6 h (80%, n = 10), 12 h (36%, n = 25), 18 h (71%, n = 24), 24 h (50%, n = 16) and 30 h (6%, n = 18) after fusion. Subsequent culture results of these cloned embryos (Table 1) indicated that there were significantly higher cleavage rates and blastocyst development in Treatment A than in Treatment B. This study clearly demonstrated that PCC is not essential to support bovine cloned blastocyst development. Direct exposure of donor nuclei in a MII cytoplast enviroment for a very short time was sufficient for nuclear reprogramming.

Table 1.
Effect of donor nuclear exposure duration in MII cytoplasm on the development potential of cloned bovine embryos
75 RABBIT NUCLEAR TRANSFER WITH CULTURED SOMATIC CELLS
Authors: Yang F. , Kessler B. , Ewerling S. , Wolf E. , Zakhartchenko V.
Published: December 2004

Cloned rabbits have been obtained by somatic cell nuclear transfer (SCNT) only with fresh, non-cultured cumulus cells (Chesne et al. 2002 Nat. Biotechnol. 20, 366–369). For the purpose of generating transgenic animals by SCNT, donor cells must be cultured and modified prior to use as nuclear donors. The objective of this study was to optimize the SCNT procedure using cultured cumulus or fibroblast cells. MII oocytes were harvested from superovulated Zika rabbits, and maternal chromosomes were removed by demecolcine-assisted enucleation (Yin et al. 2002 Biol. Reprod. 67, 442–446). Two types of somatic cells originating from Ali/Bass rabbits were used as nuclear donors: cumulus cells collected from in vivo-matured oocytes and cultured for 1–5 passages, and primary fetal fibroblasts obtained from Day 16 fetuses and grown to confluence or starved for 4–5 days. Somatic donor cells and recipient cytoplasts were fused with 2 electric pulses (1.95 kV/cm, 25 μs each, 1 s interval). Twenty to 40 min after fusion, cloned embryos were activated first with the same electropulses as for fusion, and then immediately followed by 1 h incubation in 2 mM 6-dimethylaminopurine and 5 μg/mL cytochalasin B in culture medium (B2 medium supplemented with 10% FCS). Cloned embryos were either transferred at the 2- and 4-cell stage to asynchronized recipients or cultured in vitro for 6 days. Data were compared using chi-square test, and differences were considered significant when P < 0.05. Our results demonstrate that cloned rabbits can be produced by SCNT with cultured cells but the efficiency of this technique is still very low irrespective of the type of donor cells.

Table 1.
Development of cloned embryos derived from somatic cells

This research was supported by the Therapeutic Human Polyclonals, Inc.
76 PRODUCTION OF CLONED BOVINE TRANSGENIC EMBRYOS WITH VARIOUS TYPES OF MONO-COLONY CELLS AND OVUM PICKUP
Authors: Zhao J.G. , Yang X.Y. , Huang Y. , Liu H.F. , Li H. , Li H.W. , Huang S.Z. , Zeng Y.T.
Published: December 2004

The objective of this study was to determine the effects of genetic manipulation, cell type, and culture conditions on developmental potential of bovine nuclear transfer (NT) embryos. Ovum pickup (OPU) technology was developed to obtain the oocytes for NT. A total 4044 cumulus-oocyte complexes (COCs) were obtained during 492 OPU sessions, with an average of 8.2 COCs recovered each session. Cultured granulosa cells (CGC), bovine fetal (150 days) oviduct epidermic cells (FOEC), and adult ear skin fibroblasts (ASFC) were used as donor cells for NT and were transfected with the expression vector including human FIX coding sequence directed by goat β-casein promoter and neomycin gene. The cells were screened under 800 μg mL−1 G418 for 10–14 days until the apperance of a “mono-colony” of cells which were then picked. Each cell population was expanded by consecutive passage culture under 300 μg mL−1 G418 until used for NT, ensuring that the majority of cells were transgenic. Oocytes were enucleated at 20 h post-maturation and a single donor cell was transferred into the perivitelline space of a recipient oocyte. After fusion and activation, the reconstructed embryos were co-cultured with vero cells in B2 medium for 7 days. NT efficiency between primary granulosa cells (PGC) without in vitro culture and CGC, as well as among CGC, FOEC and ASFC that were transfected with exogenous DNA (named TCGC, TFOEC, TASFC, respectively), were compared (Table 1). Differences between groups were verified by chi-square test using SAS 6.12 (SAS Institute, Inc., Cary, NC, USA) program. CGCs presented a higher fusion rate (P < 0.01) for reconstructed embryos and higher development to the blastocyst stage for NT embryos than did PGC (67% vs. 54% and 41% vs. 21%, respectively). There were no significant differences (P > 0.05) in cleavage rate (65%, 71%, and 69%, respectively) and development to the blastocyst stage for NT embryos (36%, 30% and 40%, respectively) for TCGC, TFOEC, and TASFC. A total of 86 blastocysts were selected for transfer into uteri of 86 cows, resulting in 26 pregnancies (30%) at 60 days by ultrasound scanning. Among these, 12 cows remain pregnant and 14 have aborted. The results indicated that oocytes recovered from OPU can be successfully used for NT with development to the blasocyst stage. PGC, CGC, FOEC, and ASFC can all be used for generating transgenic cattle by NT, although this needs to be verified by the birth of live calves.

Table 1.
Nuclear transfer efficiency with various cell types

This work was supported by the Chinese “863” High-Tech Plan Program (Grant No. 2002AA206201).
92 BLASTOCYST FORMATION FROM VITRIFIED BOVINE OOCYTES, ZYGOTES, AND TWO-CELL EMBRYOS
Authors: Moisan A. , Chamberlain E. , Leibo S. , Dresser B. , Bondioli K. , Godke R.
Published: December 2004

The objective of this study was to devise a protocol to preserve bovine oocytes and early cleavage-stage embryos by vitrification and to compare their subsequent embryonic development after in vitro fertilization (IVF). Mature bovine oocytes from a commercial source (BoMed; Madison, WI, USA) were randomly allocated (in four replicates) to four treatment groups. Group I: control oocytes were subjected to IVF and cultured in CR1aa medium in a humidified atmosphere of 5% O2/5% CO2/90% N2 at 38°C. Group II: MII-stage oocytes were subjected to vitrification and then fertilized by IVF. Group III: presumptive zygotes were vitrified after IVF. Group IV: two-cell embryos resulting from IVF that were cultured for ∼28 h before vitrification. The vitrification solution consisted of TCM199 medium supplemented with 10% fetal bovine serum (mTCM) and containing 20% ethylene glycol (EG)/20% dimethyl sulfoxide (DMSO)/0.65 M trehalose. The oocytes/embryos to be vitrified were rinsed in mTCM, then in 5% EG/5% DMSO, then in 10% EG/10% DMSO, and finally for 45 s in the vitrification solution. For vitrification, groups of 6 to 12 oocytes/embryos were pipetted in <1-μL volume of vitrification medium onto the tip of a CryoTop (Katayama et al. 2003 Fertil. Steril. 80, 223); plunged directly into liquid nitrogen (LN2), and stored for ∼2 h. Vitrified samples were warmed and liquefied by rapidly transferring the Cryotops from LN2 into 0.25 M trehalose in mTCM at 37°C and then sequentially at 1-min intervals into 0.188 M and 0.125 M trehalose. Cleavage was evaluated on Day 3 post-insemination, and blastocyst development was assessed on Days 7 and 9 post-insemination. Of the 251 oocytes in Group I, 71% cleaved by Day 3, 21% formed blastocysts by Day 7, and 29% did so by Day 9; 3% of the total hatched. Of the 116 oocytes in Group II, fewer cleaved (P > 0.05) by Day 3 (54%) and developed into blastocysts by Day 7 (4%) and by Day 9 (8%); none hatched. Group III zygotes (n = 131) responded like Group II oocytes, 53% cleaved, and 5% formed blastocysts on Day 7 and 7% on Day 9; none hatched. In contrast, 19% of the 122 two-cell embryos formed blastocysts by Day 7 and 28% by Day 9, and 3% hatched. Although significantly fewer oocytes/embryos in Groups II and III cleaved compared with Group I, more than 50% of them did so after vitrification. After fertilization and cleavage, the two-cell embryos were much more resistant to the deleterious effects of cryoprotectants and vitrification. Higher survival of two-cell embryos may result from their increased permeability to cryoprotectants, and to water due to their higher surface area to volume ratio.
Simulation of water and solute transport with MACRO model in Cecil loamy sand soil
Authors: Merdun Hasan , Quisenberry Virgil L.
Published: December 2004

Modelling preferential flow increases understanding of flow and transport processes in the unsaturated (vadose) zone and, hence, helps prevention of groundwater contamination. A dual-porosity model, MACRO, was evaluated for long-term drainage flow and short-term chloride-tagged water flow simulations in well-structured Cecil loamy sand soil. Water flow in micropores is calculated by the Richards’ equation, and simple gravity flow is assumed in the macropores. Solute transport in the micropores is calculated by the convection–dispersion equation (CDE), and the dispersion and diffusion in the CDE is neglected for the solute transport in the macropores. Based on the statistical criteria, the model accurately simulated drainage flow with depth and time. The average values of 3 statistical parameters (coefficient of residual mass, model efficiency, correlation coefficient) for drainage flow of different plots and times were 0.0057, 0.972, and 0.987, respectively. Similarly, the average values of the 3 statistical parameters in the same order for water flow and chloride transport of 4 plots were 0.097, 0.628, and 0.915, and 0.167, 0.938, and 0.982, respectively. The model simulated long-term drainage flow better than the short-term applied chloride-tagged water. The percentage recovery of measured water and chloride 2 h after the application ceased was 83 and 63 in the 1.05-m-deep profile of plot 1. This was a strong indication of preferential flow in this soil. Two-domain flow concept was required for acceptable simulation of this type of flow. In the 2-domain flow, boundary tension, boundary hydraulic conductivity, and effective diffusion path-length were the 3 most important parameters controlling flow and transport between the 2 domains. The effective diffusion path-length represented the structural development with depth in Cecil loamy sand soil. The relationships between the variability in flow and transport characteristics and fundamental soil properties and, hence, the associated key model parameters suggest that pedotransfer functions can be developed for the estimation of dual-porosity model parameters that control preferential flow.
Effects of temperature and application rate of a nitrification inhibitor, dicyandiamide (DCD), on nitrification rate and microbial biomass in a grazed pasture soil
Authors: Di H. J. , Cameron K. C.
Published: December 2004

Abstract. The nitrification inhibitor dicyandiamide (DCD) has recently been shown to be effective in reducing nitrate leaching from grazed pasture soils. The objective of this study was to determine the influence of temperature and application rate on the effectiveness of DCD in nitrification inhibition. Possible effects on soil microbial biomass were also determined. The soil, Lismore silt loam (Pallic orthic brown soil; Udic Haplustept loamy skeletal), was incubated at a moisture content near field capacity under 2 temperatures (8 or 20°C). Urea was applied at 25 kg N/ha and dairy cow urine at 1000 kg N/ha. DCD was applied at 2 rates equivalent to 7.5 or 15 kg/ha. The results show that at a soil temperature of 8°C, the half-life of DCD was 111–116 days. The half-life of NH4+ changed from 44 days without DCD to 243–491 days when DCD was applied. In contrast, at a soil temperature of 20°C the half-life of DCD was 18–25 days. The half-life of NH4+ changed from 22 days without DCD to 64–55 days with DCD. The 2 different rates of DCD had a small effect on the NH4+ concentration in the soil. The application of DCD did not have a significant effect on soil microbial biomass. DCD would therefore be most effective in inhibiting nitrification and thus reducing nitrate leaching in late autumn–winter–early spring in most parts of New Zealand when daily average soil temperatures are generally below 10°C and when drainage is high.
Effect of a controlled burn on the thermophysical properties of a dry soil using a new model of soil heat flow and a new high temperature heat flux sensor
Authors: Massman W. J. , Frank J. M.
Published: December 2004

Some fires can be beneficial to soils but, if a fire is sufficiently intense, soil can be irreversible altered. We measured soil temperatures and heat fluxes at several soil depths before, during, and after a controlled surface burn at Manitou Experimental Forest (southern Colorado, USA) to evaluate its effects on the soil’s thermophysical properties (thermal conductivity and volumetric specific heat capacity). During the burn the soil was heated to over 400°C at a depth of 0.02 m and to almost 100°C at 0.30 m. Relatively high temperatures persisted for several hours to days even over 1 m deep into the soil. At these intensities and durations significant changes in soil chemistry, structure, and nutrient cycling are likely. However, soil thermophysical properties, estimated before and after the fire with a new model of periodic heat flow in soils, were not significantly changed between the times shortly after sensor installation (October 2001) and 1 month after the fire (February 2002). Estimates of the soil thermophysical properties derived with the new model underestimate laboratory analyses performed on soil samples obtained after the fire. Also presented in this study are some of the first soil heat flux measurements made during a surface fire. Furthermore, data and analyses of the type discussed in this study should aid modeling studies of the soil thermal pulse associated with fire. The ultimate goal of this experiment was to provide tools to assist land managers in the use of prescribed fire to benefit ecosystems and to reduce the potential for harm by examining how the soil’s physical properties and different fuel amounts, geometries, and loading densities influence soil recovery and forest regeneration after fires.
Resolution of a paradox: Hummingbird flight at high elevation does not come without a cost
Authors: Altshuler Douglas L. , Dudley Robert , McGuire Jimmy A.
Published: December 2004

Flight at high elevation is energetically demanding because of parallel reductions in air density and oxygen availability. The hovering flight of hummingbirds is one of the most energetically expensive forms of animal locomotion, but hummingbirds are nonetheless abundant at high elevations throughout the Americas. Two mechanisms enhance aerodynamic performance in high-elevation hummingbirds: increase in wing size and wing stroke amplitude during hovering. How do these changes in morphology, kinematics, and physical properties of air combine to influence the aerodynamic power requirements of flight across elevations? Here, we present data on the flight performance of 43 Andean hummingbird species as well as a 76-taxon multilocus molecular phylogeny that served as the historical framework for comparative analyses. Along a 4,000-m elevational transect, hummingbird body mass increased systematically, placing further aerodynamic demands on high-elevation taxa. However, we found that the minimum power requirements for hovering flight remain constant with respect to elevation because hummingbirds compensate sufficiently through increases in wing size and stroke amplitude. Thus, high-elevation hummingbirds are not limited in their capacity for hovering flight despite the challenges imposed by hypobaric environments. Other flight modes including vertical ascent and fast forward flight are more mechanically and energetically demanding, and we accordingly also tested for the maximum power available to hummingbirds by using a load-lifting assay. In contrast to hovering, excess power availability decreased substantially across elevations, thereby reducing the biomechanical potential for more complex flight such as competitive and escape maneuvers.
Ancestral role of
caudal
genes in axis elongation and segmentation
Authors: Copf Tijana , Schröder Reinhard , Averof Michalis
Published: December 2004

caudal
(
cad
/
Cdx
) genes are essential for the formation of posterior structures in
Drosophila
,
Caenorhabditis elegans
, and vertebrates. In contrast to
Drosophila
, the majority of arthropods generate their segments sequentially from a posteriorly located growth zone, a process known as short-germ development.
caudal
homologues are expressed in the growth zone of diverse short-germ arthropods, but until now their functional role in these animals had not been studied. Here, we use RNA interference to examine the function of
caudal
genes in two short-germ arthropods, the crustacean
Artemia franciscana
and the beetle
Tribolium castaneum
. We show that, in both species,
caudal
is required for the formation of most body segments. In animals with reduced levels of
caudal
expression, axis elongation stops, resulting in severe truncations that remove most trunk segments. We also show that
caudal
function is required for the early phases of segmentation and Hox gene expression. The observed phenotypes suggest that in arthropods
caudal
had an ancestral role in axis elongation and segmentation, and was required for the formation of most body segments. Similarities to the function of vertebrate
Cdx
genes in the presomitic mesoderm, from which somites are generated, indicate that this role may also predate the origin of the Bilateria.
Caspases and nitric oxide broadly regulate dendritic cell maturation and surface expression of class II MHC proteins
Authors: Wong Siew Heng , Santambrogio Laura , Strominger Jack L.
Published: December 2004

The passage of dendritic cells (DC) from immature to terminally differentiated antigen-presenting cells is accompanied by numerous morphological, phenotypic, and functional changes. These changes include, for example, expression of “empty” class II MHC proteins (MHCII) at the surface in immature DC, whereas a much larger amount of peptide-loaded MHCII is expressed at the surface in mature DC. Here we show that, in cultured immature DC derived from murine bone-marrow precursors, a number of molecules involved in intracellular trafficking were present in a cleaved form, degraded by caspase-like proteases. Cleavage was either inhibited or reduced significantly during maturation of DC induced by either LPS and TNF-α or by peptides that inhibit caspase activities. Inducible nitric oxide (NO) synthetase up-regulated by LPS was essential for inhibiting the caspase-like activity during the maturation of DC. Moreover, treatment with LPS or caspase inhibitor resulted in expression of MHCII/peptide complexes at the cell surface. Thus, the alteration of the endosomal trafficking pathways during the development of DC that parallels the changes in surface expression of MHCII is regulated at least in part by the activities of caspases, inducible NO synthetase, and its product NO.
Atomic force microscopic analysis of the binding of the
Schizosaccharomyces pombe
origin recognition complex and the spOrc4 protein with origin DNA
Authors: Gaczynska Maria , Osmulski Pawel A. , Jiang Yun , Lee Joon-Kyu , Bermudez Vladimir , Hurwitz Jerard
Published: December 2004

In eukaryotes, the initiation of DNA replication requires the interaction between origin sequences and the origin recognition complex (ORC), which is highly conserved. In this report, atomic force microscopy (AFM) was used to examine the binding of
Schizosaccharomyces pombe
(sp) ORC and the spOrc4 protein with the sp autonomously replicating sequence 1 (
ars1
). AFM imaging revealed that spORC binding to
ars1
occurred solely through spOrc4p and depended on the N-terminal AT-hook domains present in spOrc4p. At high molar ratios of spORC (or spOrc4p alone) to DNA (6:1), all of the input
ars1
was bound in a one protein complex to one plasmid manner. Restriction digestion and AFM analysis of protein–DNA fragments revealed the presence of two binding sites in
ars1
. One site mapped to a region centered at nucleotide 838 of
ars1
previously detected by DNase I protection that was reported to be essential for the autonomously replicating sequence activity of
ars1
. The second site mapped to a previously uncharacterized region centered at nucleotide 1148. AFM showed that the length of the DNA fragment complexed with either spORC or spOrc4p was shortened by ≈140 bp, suggesting the wrapping of two turns of the DNA around the spOrc4p alone as well as the spOrc4p in spORC. We also show that treatment of the spORC (spOrc4p)–
ars1
complex with topoisomerase I induced a negative shift in the topoisomer distribution. These findings suggest that the binding of spORC to origin DNA alters the structure of the DNA. Thus, in the case of spORC, due to its unusual spOrc4p, at least two factors are likely to influence
ars1
activation. These include the selective binding of the complex to A- and T-rich regions and the alteration of the DNA structure due to its wrapping around spOrc4p.
Bone mass in Chinese premenarcheal girls: the roles of body composition, calcium intake and physical activity
Authors: Zhu Kun , Du Xueqin , Greenfield Heather , Zhang Qian , Ma Guansheng , Hu Xiaoqi , Fraser David R.
Published: December 2004

The association of growth and anthropometric characteristics and lifestyle factors with bone mass and second metacarpal radiogrammetry parameters was evaluated in 373 healthy Chinese premenarcheal girls aged 9–11 years. Bone mineral content (BMC) and density (BMD) and bone area (BA) of distal forearm, proximal forearm and total body, bone mineral-free lean (BMFL) mass and fat mass were measured by dual-energy X-ray absorptiometry. Metacarpal bone periosteal and medullary diameters were measured. Dietary intakes were assessed by 7d food record and physical activity (PA) by questionnaire. BMFL and fat mass together explained 6·3 and 51·6% of the variation in total body BMC and BMD, respectively. BMFL mass contributed to a substantial proportion of the variation in forearm BMC and BMD and periosteal diameter (10·4–41·0%). The corresponding BA explained 14·8–80·4% of the variation in BMC. Other minor but significant predictors of total body bone mass were Ca intake, height, age and PA score (BMD only), and of forearm bone mass were PA score, bone age, height and fat mass. Nevertheless, after adjusting for bone and body size and for age or bone age, subjects with Ca intake above the median (417mg/d) had 1·8% greater total body BMC (P<0·001), and subjects with PA scores above the median had 2·4–2·5% greater distal and proximal forearm BMC (P<0·05) than those below. Vitamin D intake negatively associated with medullary diameter (partialR21·7%). The results indicate that premenarcheal girls should be encouraged to optimise nutrition and Ca intake and exercise regularly to achieve maximum peak bone mass.
Moderate whisky consumption in combination with an evening meal reduces tryptophan availability to the brain but does not influence performance in healthy volunteers
Authors: Markus C. Rob , Sierksma Aafje , Verbeek Cees , van Rooijen Jan J. M. , Patel Hamina J. , Brand A. Nico , Hendriks Henk F. J.
Published: December 2004

Brain serotonin (5-HT) synthesis is controlled by nutrients that influence the availability of plasma tryptophan (Trp) as compared with the sum of the other large neutral amino acids (LNAA; Trp:LNAA). Alcohol consumption is found to change mood and performance and this might well be due to alterations in the plasma Trp:LNAA ratio and brain 5-HT. In the present study, we tested whether whisky consumption as part of a meal may alter the plasma Trp:LNAA ratio and influence mood and performance in healthy volunteers. Twenty-four healthy male subjects participated in a within-subjects cross-over study. Subjects consumed whisky (125ml; 40g alcohol) or water (125ml) as part of a standard evening meal. Effects of whisky consumption were tested on mood and choice reaction time and blood samples were taken to measure changes in plasma amino acids, glucose and insulin. The plasma Trp:LNAA ratio showed a significant decline 2h after whisky consumption of alcohol (P<0·001). No effects were found on choice reaction time or mood as compared with the control condition. The present findings reveal that whisky consumption alters available plasma Trp for uptake into the brain, whereas there were no effects on mood and performance.
Bioelectrical impedance phase angle as a prognostic indicator in advanced pancreatic cancer
Authors: Gupta Digant , Lis Christopher G. , Dahlk Sadie L. , Vashi Pankaj G. , Grutsch James F. , Lammersfeld Carolyn A.
Published: December 2004

Bioelectrical impedance analysis (BIA) is an easy-to-use, non-invasive and reproducible technique to evaluate changes in body composition and nutritional status. Phase angle, determined by BIA, has been found to be a prognostic indicator in several chronic conditions, such as HIV, liver cirrhosis, chronic obstructive pulmonary disease and lung cancer, and in patients undergoing dialysis. The present study investigated the prognostic role of phase angle in advanced pancreatic cancer. We evaluated a case series of fifty-eight stage IV pancreatic cancer patients treated at Cancer Treatment Centers of America®at Midwestern Regional Medical Center (Zion, IL, USA) between January 2000 and July 2003. BIA was conducted on all patients using a bioelectrical impedance analyser that operated at 50kHz. The phase angle was calculated as capacitance (Xc)/resistance (R) and expressed in degrees. The Kaplan–Meier method was used to calculate survival. Cox proportional hazard models were constructed to evaluate the prognostic effect of phase angle independent of other clinical and nutritional variables. The correlations between phase angle and traditional nutritional measures were evaluated using Pearson and Spearman coefficients. Patients with phase angle <5·0° had a median survival time of 6·3 (95% CI 3·5, 9·2) months (n29), while those with phase angle >5·0° had a median survival time of 10·2 (95% CI 9·6, 10·8) months (n29); this difference was statistically significant (P=0·02). The present study demonstrates that phase angle is a strong prognostic indicator in advanced pancreatic cancer. Similar studies in other cancer settings with larger sample sizes are needed to further validate the prognostic significance of the phase angle.
An overview of interactions between micronutrients and of micronutrients with drugs, genes and immune mechanisms
Authors: Thurnham David I.
Published: December 2004

The objective of the present review is to examine critically the consequences of interactions that micronutrients undergo with nutrients and non-nutrients (mainly prescribed medicines) in diets and lifestyle factors (smoking, tea and alcohol consumption). In addition, the review describes recent work on interactions between nutrients and genes, the influence of gene polymorphisms on micronutrients, the impact of immune responses on micronutrients and specific interactions of antioxidant micronutrients in disease processes to minimise potential pro-oxidant damage.
The key role of nutrition in controlling human population dynamics
Authors: Duncan C.J , Scott S
Published: December 2004

The early hominids and their successors, the nomadic hunter–gatherers, were evolutionarily adapted to an omnivorous diet. Their food was well balanced nutritionally and they acquired adequate supplies with relatively little expenditure of energy. The complete change to a fixed agricultural lifestyle (the Neolithic revolution) took place only some 12 000 years ago and was the most momentous event in human history. Being tied to the land that they worked led eventually to the city states and the great civilisations of history, which brought with them wars and epidemics of infectious diseases. Much more serious were the insidious effects of the new cereal-based diet which persisted until the twentieth century. Not only was it labour intensive, but also for the bulk of the population it was often deficient in vitamins, minerals and energy, particularly at certain times of the year. Time-series analysis reveals a regular short wavelength oscillation in the grain supply that persisted for at least 350 years and dominated the population dynamics of pre-industrial England. In addition to reducing fertility, it acted primarily via its effects on the nutrition of the pregnant woman. Malnutrition during one of the critical trimesters of pregnancy could have far-reaching effects not only on the health of the fetus and neonate but also on the illnesses of later, adult life. These consequences were insidiously and inevitably carried forward to the subsequent generations. Girls who were born with a low birth weight produced daughters and granddaughters of low birth weight, irrespective of their nutrition during childhood. These intergenerational, knock-on effects established a vicious circle from which there was little chance of escape.
Specific deletion of focal adhesion kinase suppresses tumor formation and blocks malignant progression
Authors: McLean Gordon W. , Komiyama Noboru H. , Serrels Bryan , Asano Hidefumi , Reynolds Louise , Conti Francesco , Hodivala-Dilke Kairbaan , Metzger Daniel , Chambon Pierre , Grant Seth G.N. , Frame Margaret C.
Published: December 2004

We have generated mice with a floxed fak allele under the control of keratin-14-driven Cre fused to a modified estrogen receptor (CreERT2). 4-Hydroxy-tamoxifen treatment induced fak deletion in the epidermis, and suppressed chemically induced skin tumor formation. Loss of fak induced once benign tumors had formed inhibited malignant progression. Although fak deletion was associated with reduced migration of keratinocytes in vitro, we found no effect on wound re-epithelialization in vivo. However, increased keratinocyte cell death was observed after fak deletion in vitro and in vivo. Our work provides the first experimental proof implicating FAK in tumorigenesis, and this is associated with enhanced apoptosis.
HHV‐8/KSHV during the development of Kaposi's sarcoma: evaluation by polymerase chain reaction and immunohistochemistry
Authors: Pak Fatemeh , Pyakural Pawan , Kokhaei Parviz , Kaaya Ephata , Pourfathollah A. A. , Selivanova Galina , Biberfeld Peter
Published: December 2004

The human γ‐herpes virus‐8 (HHV‐8) was first described in AIDS‐related Kaposi's sarcoma (KS) tumour samples. In this study, we report comparative studies on paraffin‐embedded biopsies of AIDS‐related KS (AKS) and endemic KS (EKS) with regard to HHV‐8 content as evaluated using polymerase chain reaction (PCR) and immunohistochemistry. DNA was extracted either using Chelex‐100 or using Qia‐gene kit and was evaluated with the help of a semiquantitative PCR assay. The PCR detection of HHV‐8 was more sensitive to the Chelex method than to Qia‐gene. The threshold for PCR test sensitivity with the help of serial dilution of DNA was at the level of five plasmid ORF‐26 regions, and DNA from 25 body cavity‐based lymphoma‐1 cells. The results expressed as virus load/actin unit showed progressively higher HHV‐8 levels in late (nodular) cases, compared to those in early (patch/plaque) stages. Evaluation of HHV‐8 DNA levels in tumour tissues, thus, indicates a correlation between virus load and KS stage. Double immunostaining of spindle cells (SC) in KS biopsies for CD34 and HHV‐8/latency‐associated nuclear antigen (LANA) showed an increase in double‐positive SC in the lesions of nodular AKS and EKS cases, compared to that in plaque and patch stages. However, 10–15% of CD34+/LANA– SC cells were observed during the development from patch to nodular cases of AKS and EKS. Our results indicate that PCR analysis is a simple and sensitive diagnostic method for HHV‐8 evaluation in KS tissues, processed for conventional histopathology.
Molecular diagnosis of basal cell carcinoma and other basaloid cell neoplasms of the skin by the quantification of Gli1 transcript levels
Authors: Hatta Naohito , Hirano Takashi , Kimura Tetsunori , Hashimoto Ken , Mehregan Darius R. , Ansai Shinichi , Takehara Kazuhiko , Takata Minoru
Published: December 2004

Background:  Distinguishing basal cell carcinoma (BCC) from other benign and malignant skin tumors is sometimes a difficult task for the pathologists. Because the activation of hedgehog signals and the up‐regulation of its critical transcriptional factor Gli1 are well documented in BCC, a molecular technique measuring Gli1 transcripts may aide the diagnosis.

Methods:  Gli1 transcript levels were measured by real‐time quantitative reverse transcriptase polymerase chain reaction (RT‐PCR) using RNA extracted from formalin‐fixed, paraffin‐embedded tissues of 68 cases of various skin tumors. Hematoxylin and eosin‐stained pathology slides were independently reviewed by three expert dermatopathologists.

Results:  The histological diagnoses were unambiguous in 53 tumors. The tumors included BCC (21), squamous cell carcinoma (13), seborrheic keratoses (8), trichoepithelioma (5), eccrine poroma/porocarcinoma (4), and sebaceous epithelioma/carcinoma (2). In these unambiguous cases, all BCC and trichoepithelioma tumors showed high expression of Gli1mRNA, while the expression was virtually absent in other tumors. The diagnosis was discordant among three pathologists in the remaining 15 tumors. Histological diagnoses included BCC, BCC with sebaceous differentiation, sebaceoma/sebaceous epithelioma, trichoblastoma, trichoepithelioma, basaloid follicular harmartoma, basosquamous carcinoma, etc. Six of them showed high Gli1 transcript levels.

Conclusions:  Quantification of Gli1 transcripts by RT‐PCR is helpful in discriminating BCC and trichoepithelioma from other skin tumors.
Abnormal accumulation of inter‐α‐trypsin inhibitor and hyaluronic acid in lichen sclerosus
Authors: Kuroda Kei , Fujimoto Norihiro , Tajima Shingo
Published: December 2004

Inter‐α‐trypsin inhibitor (ITI) is a recently identified extracellular hyaluronic acid (HA)‐binding protein which greatly improves extracellular HA stability. In lichen sclerosus (LS), a broad hyalinized zone of superficial dermis is the most prominent pathological change. To assess the pathogenic role of ITI in accumulation of HA in a broad hyalinized zone in LS, we examined the expression and localization of ITI and HA immunohistochemically. In LS lesional skin sections, ITI staining revealed a strong, diffuse immunoreactivity predominantly in the upper dermis, whereas no staining was detected in normal skin sections. HA staining clearly showed positive reactivity in the superficial dermis, the epidermis, and occasionally the perivascular inflammatory infiltrate in LS skin sections. In normal skin, HA was present only in the epidermis. Double staining for ITI and HA demonstrated that ITI was localized in the areas where HA was abnormally deposited in the superficial dermis of LS. Other HA‐binding proteins, CD44 and versican, did not show enhanced staining in the upper dermis of LS compared to normal skin specimens. These findings strongly suggest that ITI is closely implicated in the accumulation of HA in a broad hyalinized zone of the superficial dermis of LS.
Biphasic sarcomatoid basal cell carcinoma (carcinosarcoma): four cases with immunohistochemistry and review of the literature
Authors: Bigby S. M. , Charlton A. , Miller M. V. , Zwi L. J. , Oliver G. F.
Published: December 2004

Background:  Biphasic sarcomatoid carcinoma (BSC), or carcinosarcoma, is an uncommon biphasic neoplasm that has been reported in diverse anatomical sites. The tumor is composed of a malignant epithelial component intimately associated with a malignant mesenchymal component, which may be homologous or heterologous. Twenty‐three cases of primary cutaneous BSC have been reported in the English literature. In only eight of these cases was basal cell carcinoma the epithelial component.

Methods:  We report a further four cases of primary cutaneous biphasic basal cell carcinoma, and include the clinical, histological and immunohistochemical features.

Results:  The four cases showed basal cell carcinoma associated with a pleomorphic sarcomatous stroma. In addition, myofibroblastic differentiation and foci of osteoid were present in one case, and leiomyosarcomatous areas in another. The epithelial components were positive for several epithelial markers. The mesenchymal components were positive for vimentin and CD99, and negative for epithelial markers. p53 was positive with equal intensity in both epithelial and mesenchymal components. A significantly worse outcome was observed in patients with tumors measuring 40 mm or more at excision.

Conclusions:  The sarcomatous component of the tumor is best regarded as a metaplastic transformation of the carcinomatous component. These tumors are potentially aggressive if incompletely excised, and complete resection is recommended.
Diffuse Dermal Angiomatosis
Authors: Draper B. , Boyd A
Published: December 2004

A 53‐year‐old Caucasian male with peripheral vascular atherosclerosis presented with a 6‐week history of violaceous, tender, plaques with focal ulcerations of the mid left lower extremity. A biopsy specimen of the left leg plaque demonstrated a diffuse proliferation of benign endothelial cells in the papillary and reticular dermis with some forming small vascular spaces, occasional extravasating erythrocytes and no evidence of cholesterol emboli or vasculitis. There were no atypical endothelial mitoses or cytologic atypia. Immunohistochemical staining using endothelium‐specific anti‐CD31 antibody was positive. The clinical and pathological findings were consistent with diffuse dermal angiomatosis. The patient had a diagnostic angiogram that revealed a 1.5‐cm, focal, 80% stenosis of the midsegment of the left external iliac artery. He underwent left external iliac artery stenting with an Omnilink 8 mm by 28 mm balloon expandable stent. Poststent films showed resolution of the stenosis. Within 10 weeks of the revascularization procedure, he had complete resolution of his cutaneous manifestations. Diffuse dermal angiomatosis is a rare manifestation of a common co‐morbid condition in our ageing population. Early intervention to correct the ischemia of peripheral vascular disease results in rapid resolution of this unique clinico‐pathologic entity.
Relax, its only a Blap!
Authors: Wagamon K. , Jaworsky C.
Published: December 2004

A 67 year old female with past history of breast cancer treated with lumpectomy and post surgical radiation presented for evaluation of new papules over the breast. She was sent by her oncologist to rule out breast cancer metastases to the skin. Punch biopsy of one of the papules showed dilated vascular spaces located in the papillary dermis lined with plump endothelial cells with hyperchromatic nuclei and a hobnail like appearance. The lumen of the vascular spaces was filled with pink amorphous eosinophilic material and mitoses were not present. The stroma contained a moderately dense lymphohistiocytic infiltrate with a variable number of plasma cells. Given these features a diagnosis of benign lymphangiomatous papules of the skin (BLAP) was made. BLAPs are well reported in both the general pathology and surgical literature but have yet to be publicized in the dermatopathological literature. It is important that dermatopathologists be aware of and able to differentiate this condition from other vascular lesions, most notably well differentiated angiosarcomas.
Moderating Effects of Rater Personality on the Relation Between Candidate Self‐Monitoring and Selection Interview Ratings
Authors: Lazar Aryeh , Kravetz Shlomo , Zinger Ahuva
Published: December 2004

The present study examined the moderating effect of rater personality – extroversion and sensitivity to others – on the relations between selection interview ratings and measures of candidate self‐monitoring (SM) and social anxiety (SA). In a real‐life military selection procedure setting in which 445 candidates and 93 raters participated, rater extroversion moderated the relation between candidate SM and selection interview ratings so that this relation was negative for raters low on extroversion and positive for raters high on extroversion. Rater extroversion was also found to moderate the negative relation between candidate SA and selection interview ratings. No support was found for the moderating effect of rater sensitivity to others. An explanation of the moderating effect of rater extroversion based on the assumption that extroversion is negatively related to critical interpersonal sensitivity was suggested.
A Critical Examination of the Handbook of Industrial, Work and Organizational Psychology by N. Anderson, D.S. Ones, H.K. Sinangil and C. Viswesvaran (eds)*
Authors: Gorriti Mikel
Published: December 2004

Books reviewed:

N. Anderson, D.S. Ones, H.K. Sinangil and C. Viswesvaran, Handbook of Industrial, Work and Organizational Psychology
Immunohistochemical characterisation of the local immune response in azoxymethane‐induced colon tumours in the BDIX inbred rat strain
Authors: KOBAEK‐LARSEN MORTEN , DIEDERICHSEN AXEL , AGGER RALF , NISSEN INGER , HOKLAND MARIANNE , ZEUTHEN JESPER , RITSKES‐HOITINGA JELMERA
Published: December 2004

The aim of the present study was to characterise the local immune response in a chemically induced colon tumour model in the rat. Elucidating the character of the immune reaction may contribute to optimizing immunotherapeutic regimens for colon carcinoma in this model. Colon cancer was induced by four weekly subcutaneous azoxymethane injections in inbred rats of the BDIX/OrlIco strain in two separate studies. Azoxymethane‐induced tumours show many similarities to spontaneously occurring human colon carcinomas with respect to histopathological appearance. In our studies, the overall inflammatory reaction of the submucosa below the tumour was evaluated in haematoxylin‐eosin‐stained tissue sections. Phenotypic characterization of leukocyte infiltration in the tumour tissue was performed by immunohistochemical staining using antibodies detecting various leukocyte subsets, i.e. T cells, natural killer cells, macrophages/monocytes, and dendritic cells. The results showed that the azoxymethane‐induced colon tumours were strongly infiltrated by macrophages. Furthermore, the tumours showed a moderate degree of infiltrating CD4‐positive cells. Very few natural killer, CD8‐positive T cells and dendritic cells (identified by the OX62 antibody) were seen in the tumour tissue. Virtually no CD25‐positive cells were found. This immunohistochemical characterisation of the tumour‐infiltrating immune response in this rat model could form the basis for studies aimed at developing new immunotherapeutic regimens for human colon cancer.
Orbits of central force law potentials
Authors: Poole Charles P. , Safko John L. , Farach Horacio A.
Published: December 2004

Central force motion for power laws of the form F(r)=−krn for n>−3 (n not necessarily an integer) and k>0 is discussed in terms of the energy, angular momentum, and orbital radius of an arbitrary circular orbit. A proof is given that orbits for these power laws must be everywhere concave toward the origin. The only two cases where the orbits are closed, n=1 and n=−2, are discussed.