bluuebunny/crossref_metadata_2025 huggingface dataset:
Carotid Stiffness Indicates Risk of Ischemic Stroke and TIA in Patients With Internal Carotid Artery Stenosis
Authors: Dijk Joke M. , van der Graaf Yolanda , Grobbee Diederick E. , Bots Michiel L.
Published: August 2004

Background and Purpose—

Patients with a carotid artery stenosis, including those with an asymptomatic or moderate stenosis, have a considerable risk of ischemic stroke. Identification of risk factors for cerebrovascular disease in these patients may improve risk profiling and guide new treatment strategies. We cross-sectionally investigated whether carotid stiffness is associated with previous ischemic stroke or transient ischemic attack (TIA) in patients with a carotid artery stenosis of at least 50%.

Methods—

Patients were selected from the Second Manifestations of ARTerial disease (SMART) study, a cohort study among patients with manifest vascular disease or vascular risk factors. Arterial stiffness, measured as change in lumen diameter of the common carotid arteries during the cardiac cycle, forms part of the vascular screening performed at baseline. The first 420 participants with a stenosis of minimally 50% in at least 1 of the internal carotid arteries measured by duplex scanning were included in this study. Logistic regression analysis was used to determine the relation between arterial stiffness and previous ischemic stroke or TIA.

Results—

The risk of ischemic stroke or TIA in the highest quartile (stiffest arteries) relative to the lowest quartile was 2.1 (95% CI, 1.1 to 4.1). These findings were adjusted for age, sex, systolic blood pressure, minimal diameter of the carotid artery, and degree of carotid artery stenosis.

Conclusion—

In patients with a ≥50% carotid artery stenosis, increased common carotid stiffness is associated with previous ischemic stroke and TIA. Measurement of carotid stiffness may improve selection of high-risk patients eligible for carotid endarterectomy and may guide new treatment strategies.
Polymorphisms in Genes Involved in Inflammatory and Angiogenic Pathways and the Risk of Hemorrhagic Presentation of Brain Arteriovenous Malformations
Authors: Pawlikowska Ludmila , Tran Mary N. , Achrol Achal S. , McCulloch Charles E. , Ha Connie , Lind Denise L. , Hashimoto Tomoki , Zaroff Jonathan , Lawton Michael T. , Marchuk Douglas A. , Kwok Pui-Yan , Young William L.
Published: August 2004

Background and Purpose—

Accurate estimates of intracranial hemorrhage (ICH) risk in patients harboring brain arteriovenous malformation (BAVM) are needed to evaluate interventional strategies and to help guide clinical management. Identification of genetic polymorphisms associated with ICH would facilitate risk stratification in BAVM patients.

Methods—

We identified patients with BAVM and documented clinical presentation, demographic data, venous drainage pattern, and BAVM size. Patients were genotyped for 5 polymorphisms in 3 inflammatory cytokine genes, and 9 polymorphisms in 5 angiogenesis-related genes. Association of genotype with risk of hemorrhagic BAVM presentation was evaluated using logistic regression analysis.

Results—

We genotyped 180 patients with BAVM (53% female, 57% white, mean age at diagnosis 35±17 years, 41% presenting with ICH). BAVM patients homozygous for the interleukin 6 (
IL6
)–174G allele had a greater risk of ICH presentation (OR, 2.62,
P
=0.003) than
IL6
–174C carriers. In a multivariate logistic regression model,
IL6
–174G>C genotype, small BAVM size, and exclusively deep venous drainage were independent predictors of ICH presentation. A similar univariate trend was noted for the
TNF
α-308 GG genotype (
P
=0.055). The other polymorphisms genotyped were not associated with ICH.

Conclusions—

A polymorphism in the inflammatory cytokine IL6, but not polymorphisms in angiogenesis-related genes, was associated with ICH presentation of BAVM. Further studies are needed to define the role of inflammatory cytokines in the pathogenesis of BAVM hemorrhage.
Genetic and Environmental Contributions to Carotid Intima-Media Thickness and Obesity Phenotypes in the Northern Manhattan Family Study
Authors: Juo Suh-Hang Hank , Lin Hsiu-Fen , Rundek Tanja , Sabala Edison A. , Boden-Albala Bernadette , Park Naeun , Lan Min-Yu , Sacco Ralph L.
Published: August 2004

Background and Purpose—

Both carotid intima-media thickness (IMT) and obesity are independent determinants of stroke and cardiovascular disease. The prevalence of obesity is higher in Hispanics. The genetic basis of IMT and obesity has not been well-characterized in Caribbean Hispanics. The purpose of this study was to examine the genetic and environmental contributions to IMT and obesity in this population.

Methods—

The data included 440 subjects from 77 Caribbean Hispanic families. Mean IMT and maximum IMT were measured in the internal carotid artery, common carotid artery, and carotid bifurcation. The total IMT was calculated as the mean value of IMT at all segments. Obesity phenotypes included body mass index (BMI), waist circumference, waist-to-hip ratio (WHR), and skin-fold thickness. Variance component methods were used to estimate age-adjusted and sex-adjusted heritability. Bivariate analyses were conducted to test for genetic and environmental correlations between IMT and obesity.

Results—

Heritabilities for IMT ranged from 9% to 40%, with the highest for total maximum IMT and lowest for internal carotid artery maximum IMT. Heritabilities for BMI, waist circumference, WHR, and skin-fold thickness were 44%, 47%, 5%, and 36%, respectively. There were significant genetic, but not environmental, correlations between IMT and BMI, waist circumference, and skin-fold thickness. There were no genetic or environmental correlations between IMT and WHR.

Conclusions—

We found a substantial genetic contribution to IMT, BMI, waist circumference, and skin-fold thickness. Obesity and IMT may share common genetic factors. Future gene mapping studies are warranted to identify genes predisposing to IMT and obesity in this population.
Tracking CD40 signaling during germinal center development
Authors: Basso Katia , Klein Ulf , Niu Huifeng , Stolovitzky Gustavo A. , Tu Yuhai , Califano Andrea , Cattoretti Giorgio , Dalla-Favera Riccardo
Published: August 2004

Substantial evidence indicates that signaling through the CD40 receptor (CD40) is required for germinal center (GC) and memory B-cell formation. However, it is not fully understood at which stages of B-cell development the CD40 pathway is activated in vivo. To address this question, we induced CD40 signaling in human transformed GC B cells in vitro and identified a CD40 gene expression signature by DNA microarray analysis. This signature was then investigated in the gene expression profiles of normal B cells and found in pre- and post-GC B cells (naive and memory) but, surprisingly, not in GC B cells. This finding was validated in lymphoid tissues by showing that the nuclear factor-κB (NF-κB) transcription factors, which translocate to the nucleus upon CD40 stimulation, are retained in the cytoplasm in most GC B cells, indicating the absence of CD40 signaling. Nevertheless, a subset of centrocytes and B cells in the subepithelium showed nuclear staining of multiple NF-κB subunits, suggesting that a fraction of naive and memory B cells may be subject to CD40 signaling or to other signals that activate NF-κB. Together, these results show that GC expansion occurs in the absence of CD40 signaling, which may act only in the initial and final stages of the GC reaction. (Blood. 2004;104: 4088-4096)
Inducible chronic phase of myeloid leukemia with expansion of hematopoietic stem cells in a transgenic model of BCR-ABL leukemogenesis
Authors: Koschmieder Steffen , Göttgens Berthold , Zhang Pu , Iwasaki-Arai Junko , Akashi Koichi , Kutok Jeffery L. , Dayaram Tajhal , Geary Kristin , Green Anthony R. , Tenen Daniel G. , Huettner Claudia S.
Published: August 2004

To develop murine models of leukemogenesis, a series of transgenic mice expressing BCR-ABL in different hematopoietic cell subsets was generated. Here we describe targeted expression of P210 BCR-ABL in stem and progenitor cells of murine bone marrow using the tet-off system. The transactivator protein tTA was placed under the control of the murine stem cell leukemia (SCL) gene 3′ enhancer. Induction of BCR-ABL resulted in neutrophilia and leukocytosis, and the mice became moribund within 29 to 122 days. Autopsy of sick mice demonstrated splenomegaly, myeloid bone marrow hyperplasia, and extramedullary myeloid cell infiltration of multiple organs. BCR-ABL mRNA and protein were detectable in the affected organs. Fluorescence-activated cell sorter (FACS) analysis demonstrated a significant increase in mature and immature myeloid cells in bone marrow and spleen, together with increased bilineal B220+/Mac-1+ cells in the bone marrow. tTA mRNA was expressed in FACS-sorted hematopoietic stem cells expanded 26-fold after BCR-ABL induction. Thirty-one percent of the animals demonstrated a biphasic phenotype, consisting of neutrophilia and subsequent B-cell lymphoblastic disease, reminiscent of blast crisis. In summary, this mouse model recapitulates many characteristics of human chronic myeloid leukemia (CML) and may help elucidate basic leukemogenic mechanisms in CML stem cells during disease initiation and progression. (Blood. 2005;105:324-334)
Alpha phenyl-tert-butyl nitrone (PBN) protects syngeneic marrow transplant recipients from the lethal cytokine syndrome occurring after agonistic CD40 antibody administration
Authors: Gendelman Maria , Halligan Nadine , Komorowski Richard , Logan Brent , Murphy William J. , Blazar Bruce R. , Pritchard Kirkwood A. , Drobyski William R.
Published: August 2004

Administration of agonistic monoclonal antibodies or recombinant cytokines is a potential approach to enhance antitumor immunity in bone marrow (BM) transplant recipients, but is complicated by toxicity due to proinflammatory cytokine-mediated vital organ damage. We used a murine syngeneic bone marrow transplant (BMT) model, in which administration of anti-CD40 antibody early after BMT results in overproduction of interleukin-12 (IL-12) and interferon-γ (IFN-γ), and lethal gut toxicity to examine the protective effect of the spin trap inhibitor, alpha phenyl-tert-butyl nitrone (PBN). Administration of PBN protected transplant recipients from mortality by significantly attenuating gut toxicity, but did not effect a reduction in the levels of proinflammatory cytokines (IL-12, IFN-γ, tumor necrosis factor α [TNF-α], or nitrate/nitrite). Moreover, PBN did not compromise anti-CD40 antibody-mediated antitumor effects in a nontransplantation lymphoma model. Collectively, these data suggest that PBN administration may represent a novel approach for reduction of toxicity without compromise of antitumor effects resulting from administration of therapeutic antibodies in both transplantation and nontransplantation settings. (Blood. 2005;105:428-431)
MRP8 and MRP14 control microtubule reorganization during transendothelial migration of phagocytes
Authors: Vogl Thomas , Ludwig Stephan , Goebeler Matthias , Strey Anke , Thorey Irmgard S. , Reichelt Rudolf , Foell Dirk , Gerke Volker , Manitz Marie P. , Nacken Wolfgang , Werner Sabine , Sorg Clemens , Roth Johannes
Published: August 2004

MRP14 (S100A9) is the major calcium-binding protein of neutrophils and monocytes. Targeted gene disruption reveals an essential role of this S100 protein for transendothelial migration of phagocytes. The underlying molecular mechanism comprises major alterations of cytoskeletal metabolism. MRP14, in complex with its binding partner MRP8 (S100A8), promotes polymerization of microtubules. MRP14 is specifically phosphorylated by p38 mitogen-activated protein kinase (MAPK). This phosphorylation inhibits MRP8/MRP14-induced tubulin polymerization. Phosphorylation of MRP14 is antagonistically regulated by binding of MRP8 and calcium. The biologic relevance of these findings is confirmed by the fact that MAPK p38 fails to stimulate migration of MRP14-/- granulocytes in vitro and MRP14-/- mice show a diminished recruitment of granulocytes into the granulation tissue during wound healing in vivo. MRP14-/- granulocytes contain significantly less polymerized tubulin, which subsequently results in minor activation of Rac1 and Cdc42 after stimulation of p38 MAPK. Thus, the complex of MRP8/MRP14 is the first characterized molecular target integrating MAPK- and calcium-dependent signals during migration of phagocytes.
Re-establishment of VWF-dependent Weibel-Palade bodies in VWD endothelial cells
Authors: Haberichter Sandra L. , Merricks Elizabeth P. , Fahs Scot A. , Christopherson Pamela A. , Nichols Timothy C. , Montgomery Robert R.
Published: August 2004

Type 3 von Willebrand disease (VWD) is a severe hemorrhagic defect in humans. We now identify the homozygous mutation in the Chapel Hill strain of canine type 3 VWD that results in premature termination of von Willebrand factor (VWF) protein synthesis. We cultured endothelium from VWD and normal dogs to study intracellular VWF trafficking and Weibel-Palade body formation. Weibel-Palade bodies could not be identified in the canine VWD aortic endothelial cells (VWD-AECs) by P-selectin, VWFpp, or VWF immunostaining and confocal microscopy. We demonstrate the reestablishment of Weibel-Palade bodies that recruit endogenous P-selectin by expressing wild-type VWF in VWD-AECs. Expression of mutant VWF proteins confirmed that VWF multimerization is not necessary for Weibel-Palade body creation. Although the VWF propeptide is required for the formation of Weibel-Palade bodies, it cannot independently induce the formation of the granule. These VWF-null endothelial cells provide a unique opportunity to examine the biogenesis of Weibel-Palade bodies in endothelium from a canine model of type 3 VWD.
Risk of head and neck squamous cell cancer and death in patients with Fanconi anemia who did and did not receive transplants
Authors: Rosenberg Philip S. , Socié Gerard , Alter Blanche P. , Gluckman Eliane
Published: August 2004

Hematopoietic stem cell transplant (SCT) is currently the only therapy that can restore normal hematopoiesis in patients with Fanconi anemia (FA). Patients with FA have a high baseline risk of squamous cell cancers (SCCs) of the head, neck, and esophagus, and SCT conditioning may increase SCC incidence. We evaluated the risks of SCC and death in 145 patients with FA in the North American Survey (NAS) cohort who did not receive transplants, and 117 patients with FA in the Hôpital Saint Louis (SLH) cohort who did receive transplants. The age-specific hazard of SCC was 4.4-fold higher in patients who received transplants than in those who did not (P = .003), and SCCs occurred at significantly younger ages in the former (respective medians, 18 and 33 years, P = .004). Survival after SCC was similarly poor in both cohorts (P = .135, median, 13 months). The hazard of SCC increased at a greater than linear rate, to 4.4% per year by age 40 in NAS and 4.7% per year by 10 years after transplant in SLH. In SLH, the hazard of non-SCC death was biphasic, declining significantly (P = .004) from 7.1% per month during the first 6 months after transplant to 0.13% per month (1.6% per year) after the first year. Acute and chronic graft-versus-host diseases were significant SCC risk factors. Adverse event rates in these cohorts provide historical control rates to assess emerging therapies for FA.
Noninvasive MR imaging of magnetically labeled stem cells to directly identify neovasculature in a glioma model
Authors: Anderson Stasia A. , Glod John , Arbab Ali S. , Noel Martha , Ashari Parwana , Fine Howard A. , Frank Joseph A.
Published: August 2004

Bone marrow-derived endothelial precursor cells incorporate into neovasculature and have been successfully used as vehicles for gene delivery to brain tumors. To determine whether systemically administered Sca1+ bone marrow cells labeled with superparamagnetic iron oxide nanoparticles can be detected by in vivo magnetic resonance imaging in a mouse brain tumor model, mouse Sca1+ cells were labeled in vitro with ferumoxides-poly-l-lysine complexes. Labeled or control cells were administered intravenously to glioma-bearing severe combined immunodeficient (SCID) mice. Magnetic resonance imaging (MRI) was performed during tumor growth. Mice that received labeled cells demonstrated hypointense regions within the tumor that evolved over time and developed a continuous dark hypointense ring at a consistent time point. This effect was not cleared by administration of a gadolinium contrast agent. Histology showed iron-labeled cells around the tumor rim in labeled mice, which expressed CD31 and von Willebrand factor, indicating the transplanted cells detected in the tumor have differentiated into endothelial-like cells. These results demonstrate that MRI can detect the incorporation of magnetically labeled bone marrow-derived precursor cells into tumor vasculature as part of ongoing angiogenesis and neovascularization. This technique can be used to directly identify neovasculature in vivo and to facilitate gene therapy by noninvasively monitoring these cells as gene delivery vectors. (Blood. 2005;105:420-425)
A supertree approach to shorebird phylogeny
Authors: Thomas Gavin H , Wills Matthew A , Székely Tamás
Published: August 2004

Background

Order Charadriiformes (shorebirds) is an ideal model group in which to study a wide range of behavioural, ecological and macroevolutionary processes across species. However, comparative studies depend on phylogeny to control for the effects of shared evolutionary history. Although numerous hypotheses have been presented for subsets of the Charadriiformes none to date include all recognised species. Here we use the matrix representation with parsimony method to produce the first fully inclusive supertree of Charadriiformes. We also provide preliminary estimates of ages for all nodes in the tree.

Results

Three main lineages are revealed: i) the plovers and allies; ii) the gulls and allies; and iii) the sandpipers and allies. The relative position of these clades is unresolved in the strict consensus tree but a 50% majority-rule consensus tree indicates that the sandpiper clade is sister group to the gulls and allies whilst the plover group is placed at the base of the tree. The overall topology is highly consistent with recent molecular hypotheses of shorebird phylogeny.

Conclusion

The supertree hypothesis presented herein is (to our knowledge) the only complete phylogenetic hypothesis of all extant shorebirds. Despite concerns over the robustness of supertrees (see Discussion), we believe that it provides a valuable framework for testing numerous evolutionary hypotheses relating to the diversity of behaviour, ecology and life-history of the Charadriiformes.
Recurrent Barrett's esophagus and adenocarcinoma after esophagectomy
Authors: Wolfsen Herbert C , Hemminger Lois L , DeVault Kenneth R
Published: August 2004

Background

Esophagectomy is considered the gold standard for the treatment of high-grade dysplasia in Barrett's esophagus (BE) and for noninvasive adenocarcinoma (ACA) of the distal esophagus. If all of the metaplastic epithelium is removed, the patient is considered "cured". Despite this, BE has been reported in patients who have previously undergone esophagectomy. It is often debated whether this is "new" BE or the result of an esophagectomy that did not include a sufficiently proximal margin. Our aim was to determine if BE recurred in esophagectomy patients where the entire segment of BE had been removed.

Methods

Records were searched for patients who had undergone esophagectomy for cure at our institution. Records were reviewed for surgical, endoscopic, and histopathologic findings. The patients in whom we have endoscopic follow-up are the subjects of this report.

Results

Since 1995, 45 patients have undergone esophagectomy for cure for Barrett's dysplasia or localized ACA. Thirty-six of these 45 patients underwent endoscopy after surgery including 8/45 patients (18%) with recurrent Barrett's metaplasia or neoplasia after curative resection.

Conclusion

Recurrent Barrett's esophagus or adenocarcinoma after esophagectomy was common in our patients who underwent at least one endoscopy after surgery. This appears to represent the development of metachronous disease after complete resection of esophageal disease. Half of these patients have required subsequent treatment thus far, either repeat surgery or photodynamic therapy. These results support the use of endoscopic surveillance in patients who have undergone "curative" esophagectomy for Barrett's dysplasia or localized cancer.
Comparison of bovine leukemia virus (BLV) and CMV promoter-driven reporter gene expression in BLV-infected and non-infected cells
Authors: Harms Jerome S , Eakle Kurt A , Kuo Lillian S , Bremel Robert D , Splitter Gary A
Published: August 2004

Background

Viral promoters are used in mammalian expression vectors because they generally have strong activity in a wide variety of cells of differing tissues and species.

Methods

The utility of the BLV LTR/promoter (BLVp) for use in mammalian expression vectors was investigated through direct comparison to the CMV promoter (CMVp). Promoter activity was measured using luciferase assays of cell lines from different tissues and species stably transduced with BLVp or CMVp driven luciferase vectors including D17, FLK, BL3.1 and primary bovine B cells. Cells were also modified through the addition of BLV Tax expression vectors and/or BLV infection as well as treatment with trichostatin A (TSA).

Results

Results indicate the BLV promoter, while having low basal activity compared to the CMV promoter, can be induced to high-levels of activity similar to the CMV promoter in all cells tested. Tax or BLV infection specifically enhanced BLVp activity with no effect on CMVp activity. In contrast, the non-specific activator, TSA, enhanced both BLVp and CMVp activity.

Conclusion

Based on these data, we conclude the BLV promoter could be very useful for transgene expression in mammalian expression vectors.
Comprehensive analysis of pseudogenes in prokaryotes: widespread gene decay and failure of putative horizontally transferred genes
Authors: Liu Yang , Harrison Paul M , Kunin Victor , Gerstein Mark
Published: August 2004

Background

Pseudogenes often manifest themselves as disabled copies of known genes. In prokaryotes, it was generally believed (with a few well-known exceptions) that they were rare.

Results

We have carried out a comprehensive analysis of the occurrence of pseudogenes in a diverse selection of 64 prokaryote genomes. Overall, we find a total of around 7,000 candidate pseudogenes. Moreover, in all the genomes surveyed, pseudogenes occur in at least 1 to 5% of all gene-like sequences, with some genomes having considerably higher occurrence. Although many large populations of pseudogenes arise from large, diverse protein families (for example, the ABC transporters), notable numbers of pseudogenes are associated with specific families that do not occur that widely. These include the cytochrome P450 and PPE families (PF00067 and PF00823) and others that have a direct role in DNA transposition.

Conclusions

We find suggestive evidence that a large fraction of prokaryote pseudogenes arose from failed horizontal transfer events. In particular, we find that pseudogenes are more than twice as likely as genes to have anomalous codon usage associated with horizontal transfer. Moreover, we found a significant difference in the number of horizontally transferred pseudogenes in pathogenic and non-pathogenic strains of Escherichia coli.
Function-informed transcriptome analysis of Drosophila renal tubule
Authors: Wang Jing , Kean Laura , Yang Jingli , Allan Adrian K , Davies Shireen A , Herzyk Pawel , Dow Julian AT
Published: August 2004

Background

Comprehensive, tissue-specific, microarray analysis is a potent tool for the identification of tightly defined expression patterns that might be missed in whole-organism scans. We applied such an analysis to Drosophila melanogaster Malpighian (renal) tubule, a defined differentiated tissue.

Results

The transcriptome of the D. melanogaster Malpighian tubule is highly reproducible and significantly different from that obtained from whole-organism arrays. More than 200 genes are more than 10-fold enriched and over 1,000 are significantly enriched. Of the top 200 genes, only 18 have previously been named, and only 45% have even estimates of function. In addition, 30 transcription factors, not previously implicated in tubule development, are shown to be enriched in adult tubule, and their expression patterns respect precisely the domains and cell types previously identified by enhancer trapping. Of Drosophila genes with close human disease homologs, 50 are enriched threefold or more, and eight enriched 10-fold or more, in tubule. Intriguingly, several of these diseases have human renal phenotypes, implying close conservation of renal function across 400 million years of divergent evolution.

Conclusions

From those genes that are identifiable, a radically new view of the function of the tubule, emphasizing solute transport rather than fluid secretion, can be obtained. The results illustrate the phenotype gap: historically, the effort expended on a model organism has tended to concentrate on a relatively small set of processes, rather than on the spread of genes in the genome.
Improving identification of differentially expressed genes in microarray studies using information from public databases
Authors: Kim Richard D , Park Peter J
Published: August 2004

We demonstrate that the process of identifying differentially expressed genes in microarray studies with small sample sizes can be substantially improved by extracting information from a large number of datasets accumulated in public databases. The improvement comes from more reliable estimates of gene-specific variances based on other datasets. For a two-group comparison with two arrays in each group, for example, the result of our method was comparable to that of a t-test analysis with five samples in each group or to that of a regularized t-test analysis with three samples in each group. Our results are further improved by weighting the results of our approach with the regularized t-test results in a hybrid method.
Regulation of Steroidogenesis by p53 in Macaque Granulosa Cells and H295R Human Adrenocortical Cells
Authors: Cherian-Shaw Mary , Das Rituparna , VandeVoort Catherine A. , Chaffin Charles L.
Published: August 2004

Ovulation and formation of a functional corpus luteum in primates involve cascades of events, including increased progesterone synthesis and changes in granulosa cell proliferation. However, critical gaps remain in our understanding of how an ovulatory gonadotropin surge initiates these processes. To more fully elucidate changes in the cell cycle during luteal formation, the actions of the tumor suppressor p53 were examined. Rhesus macaque granulosa cells were isolated during controlled ovarian stimulation protocols before (nonluteinized) or after (luteinized) an ovulatory gonadotropin stimulus. Phosphorylated p53 protein was detected in the cytoplasm of granulosa cells before and after human chorionic gonadotropin (hCG) treatment, whereas granulosa cells from hormonally controlled rats did not express p53 before or after hCG. Treatment of nonluteinized macaque granulosa cells with hCG and the p53 inhibitor pifithrin-α (PFT) in vitro did not alter markers of the cell cycle, including proliferating cell nuclear antigen, p21, and human double minute (HDM)-2 expression compared with hCG alone. Levels of pregnenolone and progesterone increased 2- and 4-fold, respectively, within 6 h of hCG treatment, whereas PFT completely blocked this hCG-induced effect. Estradiol was increased transiently (>10-fold) by hCG plus PFT relative to levels after hCG alone. PFT also inhibited hCG-induced increases in steroidogenic acute regulatory protein and 3β-hydroxysteroid dehydrogenase mRNAs. Similar results were obtained using the human adrenocortical cell line H295R, suggesting that p53 may have a general function in primate steroidogenesis. These data indicate that p53 plays a key role in luteinization of the primate ovarian follicle though the regulation of steroidogenic enzymes leading to progesterone synthesis.
Evidence for a Novel Intrapituitary Autocrine/Paracrine Feedback Loop Regulating Growth Hormone Synthesis and Secretion in Grass Carp Pituitary Cells by Functional Interactions between Gonadotrophs and Somatotrophs
Authors: Zhou Hong , Wang Xinyan , Ko Wendy K. W. , Wong Anderson O. L.
Published: August 2004

Gonadotropin (GTH) and GH released from the pituitary are known to interact at multiple levels to modulate the functions of the gonadotrophic and somatotrophic axes. However, their interactions at the pituitary level have not been fully characterized. In this study, autocrine/paracrine regulation of GH synthesis and secretion by local interactions between gonadotrophs and somatotrophs was examined using grass carp pituitary cells as a cell model. Exogenous GTH and GH induced GH release and GH mRNA expression in carp pituitary cells. Removal of endogenous GTH and GH by immunoneutralization with GTH and GH antisera, respectively, suppressed GH release, GH production, and GH mRNA levels. GH antiserum also blocked the stimulatory effects of exogenous GTH on GH release and GH mRNA levels. In reciprocal experiments, GH release and GH mRNA expression induced by exogenous GH was significantly reduced by GTH antiserum. In addition, exogenous GH was found to be inhibitory to basal GTH release and treatment with GH antiserum elevated GTH secretion at low doses but suppressed GTH production at high doses. These results suggest that local interactions between gonadotrophs and somatotrophs may form an intrapituitary feedback loop to regulate GH release and synthesis. In this model, GTH released from gonadotrophs induces GH release and GH production in neighboring somatotrophs. GH secreted maintains somatotroph sensitivity to GTH stimulation, and at the same time, inhibits basal GTH release in gonadotrophs. This feedback loop may represent a novel mechanism regulating GH release and synthesis in lower vertebrates.
Minireview: GNAS: Normal and Abnormal Functions
Authors: Weinstein Lee S. , Liu Jie , Sakamoto Akio , Xie Tao , Chen Min
Published: August 2004

GNAS is a complex imprinted gene that uses multiple promoters to generate several gene products, including the G protein α-subunit (Gsα) that couples seven-transmembrane receptors to the cAMP-generating enzyme adenylyl cyclase. Somatic activating Gsα mutations, which alter key residues required for the GTPase turn-off reaction, are present in various endocrine tumors and fibrous dysplasia of bone, and in a more widespread distribution in patients with McCune- Albright syndrome. Heterozygous inactivating Gsα mutations lead to Albright hereditary osteodystrophy. Gsα is imprinted in a tissue-specific manner, being primarily expressed from the maternal allele in renal proximal tubules, thyroid, pituitary, and ovary. Maternally inherited mutations lead to Albright hereditary osteodystrophy (AHO) plus PTH, TSH, and gonadotropin resistance (pseudohypoparathyroidism type 1A), whereas paternally inherited mutations lead to AHO alone. Pseudohypoparathyroidism type 1B, in which patients develop PTH resistance without AHO, is almost always associated with a GNAS imprinting defect in which both alleles have a paternal-specific imprinting pattern on both parental alleles. Familial forms of the disease are associated with a mutation within a closely linked gene that deletes a region that is presumably required for establishing the maternal imprint, and therefore maternal inheritance of the mutation results in the GNAS imprinting defect. Imprinting of one differentially methylated region within GNAS is virtually always lost in pseudohypoparathyroidism type 1B, and this region is probably responsible for tissue-specific Gsα imprinting. Mouse knockout models show that Gsα and the alternative Gsα isoform XLαs that is expressed from the paternal GNAS allele may have opposite effects on energy metabolism in mice.
Minireview: PRKAR1A: Normal and Abnormal Functions
Authors: Bossis Ioannis , Stratakis Constantine A.
Published: August 2004

The type 1α regulatory subunit (RIα) of cAMP-dependent protein kinase (PKA) (coded by the PRKAR1A gene) is the main component of type I PKA, which regulates most of the serine-threonine kinase activity catalyzed by the PKA holoenzyme in response to cAMP. Carney complex (CNC), or the complex of spotty skin pigmentation, myxomas, and endocrine overactivity, is a multiple endocrine (and not only) neoplasia syndrome that is due to PRKAR1A-inactivating mutations. The R1α protein and PRKAR1A mRNA have been found to be up-regulated in a series of cell lines and human and rodent neoplasms, suggesting this molecule’s involvement in tumorigenesis and its potential role in cell cycle regulation, growth, and/or proliferation. Alterations in PKA activity elicit a variety of effects depending on the tissue, developmental stage, degree of differentiation, and cAMP levels. In addition, RIα may have functions independent of PKA. The presence of inactivating germline mutations and the loss of its wild-type allele in some CNC lesions indicate that PRKAR1A might function as a tumor suppressor gene in these tissues, but could PRKAR1A be a classic tumor suppressor gene? Probably not, and this review explains why.
Minireview: Cyclin D1: Normal and Abnormal Functions
Authors: Fu Maofu , Wang Chenguang , Li Zhiping , Sakamaki Toshiyuki , Pestell Richard G.
Published: August 2004

Cyclin D1 encodes the regulatory subunit of a holoenzyme that phosphorylates and inactivates the retinoblastoma protein and promotes progression through the G1-S phase of the cell cycle. Amplification or overexpression of cyclin D1 plays pivotal roles in the development of a subset of human cancers including parathyroid adenoma, breast cancer, colon cancer, lymphoma, melanoma, and prostate cancer. Of the three D-type cyclins, each of which binds cyclin-dependent kinase (CDK), it is cyclin D1 overexpression that is predominantly associated with human tumorigenesis and cellular metastases. In recent years accumulating evidence suggests that in addition to its original description as a CDK-dependent regulator of the cell cycle, cyclin D1 also conveys cell cycle or CDK-independent functions. Cyclin D1 associates with, and regulates activity of, transcription factors, coactivators and corepressors that govern histone acetylation and chromatin remodeling proteins. The recent findings that cyclin D1 regulates cellular metabolism, fat cell differentiation and cellular migration have refocused attention on novel functions of cyclin D1 and their possible role in tumorigenesis. In this review, both the classic and novel functions of cyclin D1 are discussed with emphasis on the CDK-independent functions of cyclin D1.
Selective Recruitment of p160 Coactivators on Glucocorticoid-Regulated Promoters in Schwann Cells
Authors: Grenier Julien , Trousson Amalia , Chauchereau Anne , Amazit Larbi , Lamirand Audrey , Leclerc Philippe , Guiochon-Mantel Anne , Schumacher Michael , Massaad Charbel
Published: August 2004

In the nervous system, glucocorticoid hormones play a major role during development and throughout life. We studied the mechanisms of action of the glucocorticoid receptor (GR) and its interactions with p160 coactivator family members [steroid receptor coactivator (SRC)-1 (a and e), SRC-2 and SRC-3] in mouse Schwann cells (MSC80). We found that the three p160s were expressed in MSC80 cells. We have shown by functional overexpression and RNA interference experiments that the recruitment of these coactivators by the GR is promoter dependent. A minimal promoter containing two glucocorticoid response elements, (GRE)2-TATA, recruits SRC-1 (a and e) and SRC-3, whereas SRC-2 is excluded. Within the context of the more complex mouse mammary tumor virus promoter, GR recruits SRC-1e and SRC-2, whereas SRC-1a and SRC-3 are not implicated. Furthermore, we have identified cytosolic aspartate aminotransferase as a GR target gene in MSC80 cells by microarray experiments. The GR recruits exclusively SRC-1e in the context of the cytosolic aspartate aminotransferase promoter. Because SRC-1 is the omnipresent coactivator of GR, we further investigated the interactions between GR and this coactivator in Schwann cells by reporter assays and immunocytochemistry experiments with deleted forms of SRC-1. We have shown that SRC-1 unexpectedly interacts with GR via its two nuclear receptor binding domains, thus providing a novel mechanism of GR signaling within the nervous system.
The Rarity of “Unusal” Dispositions of Victim Bodies: Staging and Posing
Authors: Keppel RD , Weis JG
Published: August 2004

The act of leaving a victim's body in an unusual position is a conscious criminal action by an offender to thwart an investigation, shock the finder and investigators of the crime scene, or give perverted pleasure to the killer. The unusual position concepts of posing and staging a murder victim have been documented thoroughly and have been accepted by the courts as a definable phenomenon. One staging case and one posing case are outlined and reveal characteristics of those homicides. From the Washington State Attorney General's Homicide Investigation and Tracking System's database on murder covering the years 1981–2000 (a total of 5,224 cases), the relative frequency of unusual body dispositions is revealed as a very rare occurrence. Only 1.3% of victims are left in an unusual position, with 0.3% being posed and 0.1% being staged. The characteristics of these types of murders also set them apart: compared to all other murders, in staged murders the victims and killers are, on average, older. All victims and offenders in the staged murders are white, with victims being disproportionately white in murders with any kind of unusual body disposition. Likewise, females stand out as victims when the body is posed, staged, or left in other unusual positions. Whereas posed bodies are more likely to include sexual assault, often in serial murders, there is no evidence of either in the staged cases. Lastly, when a body is left in an unusual position, binding is more likely, as well as the use of more “hands on” means of killing the victim, such as stabbing or cutting weapons, bludgeons, ligatures, or hands and feet. Erratum to this paper appears in 49(6).
A Proposed Method for the Identification of Race in Sub-Adult Skeletons: A Geometric Morphometric Analysis of Mandibular Morphology
Authors: Buck TJ , Vidarsdottir US
Published: August 2004

The identification of biological race (ancestry) in skeletal material is an important aspect of forensic investigations. While techniques for race determination are well established for adult skeletons, identification of race in sub-adult specimens has not been widely addressed. The present study investigates racial differences in the mandibular morphology of sub-adult specimens using geometric morphometric analyses. One hundred and seventy-four mandibles from five morphologically distinct samples were digitized and subjected to general Procrustes analysis. Results showed significant morphological differences between the samples and obtained cross-validation results of over 70% accuracy in identification of unknown individuals using the complete mandible. It is suggested that these techniques could provide a method for the identification of race in sub-adult individuals.
Fatality Due to Methyl Acetylene-Propadiene (MAPP) Inhalation
Authors: Avella J , Lehrer M
Published: August 2004

A 33-year-old man died after intentionally inhaling a gaseous mix of methyl acetylene (propyne) and propadiene (allene) commonly known as MAPP, which is used for soldering and welding. He was found with a plastic bag securely placed over his head and a cylinder of MAPP alongside his head. The cylinder had been vented into the bag using a flexible hose. A comprehensive toxicological analysis revealed only a trace of diphenhydramine in the liver and 0.02 mg/L of morphine in the urine. Analysis of blood by headspace gas chromatography (HS-GC) detected two unknown peaks. These were determined to be the components of MAPP gas. MAPP was quantitated in femoral blood (59.6 mg/L) and brain (43.6 mg/kg) using a HS-GC method. The cause of death was attributed to acute MAPP intoxication, and the manner was determined to be suicide. A discussion on the analytical and interpretive considerations commonly encountered when analyzing volatile compounds is also presented.
Effects of cyclic steroid hormone replacement on prolactin and luteinizing hormone surges in female rats
Authors: Brown A M , Janik J M , Murphree E S , King R , Callahan P
Published: August 2004

The ability of steroid hormones to produce an LH or prolactin (PRL) surge was determined in rats ovariectomized at 6, 9 or 13 weeks of age and subjected to one, three or six cycles of estrogen and progesterone replacement. Sensitivity to steroid replacement was dependent on the age of the animal at the time of ovariectomy. Repeated cyclic steroid hormone replacement significantly increased the magnitude of the PRL response, but not the LH response, in animals ovariectomized at 6 weeks. The LH response was significantly altered by cyclic steroid replacement only in animals ovariectomized at 13 weeks. These results indicate that the mechanisms involved in the regulation of PRL secretion are influenced by steroid hormone replacement and that cyclic steroid hormone exposure increases the magnitude of the PRL secretory response.
Changes in the reproductive system of male mice immunized with a GnRH-analogue conjugated to mycobacterial hsp70
Authors: Hannesdóttir S G , Han X , Lund T , Singh M , van der Zee R , Roitt I M , Delves P J
Published: August 2004

Immunosterilization is an attractive alternative to surgical castration. Gonadotropin-releasing hormone (GnRH) controls the production of the gonadotropins thereby having an orchestrating effect on the reproductive hormone cascade and spermatogenesis. Induction of neutralizing antibody can abrogate the effect of the hormone. Current GnRH-based vaccines often require strong adjuvants and/or multiple injections of the vaccines to overcome variability in the response. Heat shock proteins (hsp) have been used as carrier molecules because of their powerful intrinsic ability to enhance an immune response to associated antigens. A GnRH-analogue, GnRH-d6-Lys, was conjugated to recombinantMycobacterium tuberculosishsp70. Male BALB/c mice were immunized i.p. with GnRH-hsp70 in the mild adjuvant Ribi or in incomplete Freund’s adjuvant (IFA). The initial immunizations were done on pre-pubertal 3-week-old mice, with boosts at 5 and 8 weeks of age. The mice were killed at 10 weeks of age and GnRH-specific antibodies and serum testosterone levels measured. All the immunized mice mounted GnRH-specific antibody responses, with no difference in the mice immunized with GnRH-hsp70/Ribi or with GnRH-hsp70/IFA. There was substantial atrophy of the urogenital complex and significantly (P< 0.0005) reduced levels of testosterone-dependent testicular relaxin-like factor mRNA expression. Mice immunized with GnRH-hsp70/Ribi showed substantially reduced (P< 0.001) serum testosterone levels. These results indicate that hsp70 may serve as a particularly advantageous carrier for GnRH-based vaccines.
Primate models for assisted reproductive technologies
Authors: Hewitson Laura
Published: August 2004

Although the deliberate creation of human embryos for scientific research is complicated by ethical and practical issues, a detailed understanding of the cellular and molecular events occurring during human fertilization is essential, particularly for understanding infertility. It is clear from cytoskeletal imaging studies of mouse fertilization that this information cannot be extrapolated to humans because of unique differences in centrosomal inheritance. However, the cytoskeletal rearrangements during non-human primate fertilization are very similar to humans, providing a compelling animal model in which to examine sperm–egg interactions. In order to address this key step in primate fertilization and to avoid the complexities in working with fertilized human zygotes, studies are now exploring the molecular foundations of various assisted fertilization techniques in a monkey model. While intracytoplasmic sperm injection with ejaculated or testicular sperm is quite successful in primate models, there are some specific differences when compared with standard IVF that warrant further investigation, particularly in regards to nuclear remodeling, genomic imprinting, Y-chromosome deletions and developmental outcomes. Similarly, primate models have been useful for examining spermatid function during fertilization but these have met with limited success. One area of primate reproductive research that has yet to be mastered is reproductive cloning. Genetically identical primates would provide the ultimate approach for accelerating stem cell-based therapies for a number of neurodegenerative diseases such as Alzheimer’s and Parkinson’s disease, as well as targeted gene therapies for various metabolic disorders.
Linking Stress to the Increased Susceptibility of Channel Catfish to Enteric Septicemia Using Cortisol
Authors: Sink Todd D. , Strange Richard J.
Published: August 2004

Although stress is known to increase the susceptibility of channel catfish to disease, this has not been correlated to a clinical measure of stress like plasma cortisol. By exposing healthy, unstressed catfish fingerlings (70–85 mm) to six concentrations of Edwardsiella ictaluri, we determined that the dose at which 30% of the fingerlings would die (LD30) was 1.4 log colony-forming units (CFU) per liter. Twenty-five small fingerling catfish per tank were confined to 12.5-cm × 15-cm × 10-cm plastic nets for 30 or 60 min, while unstressed fish remained free-swimming. Blood samples were taken for cortisol analysis, and the LD30 dose of the pathogen was administered at the conclusion of the stress period. Unstressed fish were sampled and challenged similarly. Infection rates were monitored for 21 d. A second challenge was conducted with larger fingerlings (114–168 mm) in the same manner. In the experiment with small fingerlings, cortisol concentrations (mean ± SD) for unstressed fish (23.6 ± 6.02 ng/mL), fish stressed 30 min (77.7 ± 19.16 ng/mL), and fish stressed 60 min (115.4 ± 26.65 ng/mL) differed significantly (P < 0.05) from each other. Significant increases in mortality occurred in conjunction with increases in cortisol (unstressed: 22.5 ± 4.2%, 30 min: 47.5 ± 5.2%, 60 min: 81.7 ± 6.8%). For large fingerlings, cortisol concentrations also differed between unstressed fish (31.44 ± 17.9 ng/mL) and stressed fish (30 min: 103.1 ± 20.7 ng/mL, 60 min: 109.6 ± 24.9 ng/mL). Although cortisol did not differ notably between the 30- and 60-min stress groups, mortalities in the large fingerlings increased significantly with level of stress (unstressed: 25.8 ± 7.4%, 30 min: 62.5 ± 10.4%, 60 min: 100 ± 0.0%). This study demonstrates the relationship between cortisol and the increased susceptibility of channel catfish to enteric septicemia of catfish.
Analysis of Protein S‐Nitrosylation
Authors: Mannick Joan B. , Schonhoff Christopher M.
Published: August 2004

S‐nitrosylation is the binding of an NO group to a cysteine or other thiol. Like phosphorylation, S‐nitrosylation is a precisely targeted and rapidly reversible post‐translational modification that serves as an on/off switch for protein function during cell signaling. However, unlike phosphorylation, S‐nitrosylation of proteins occurs nonenzymatically, mediated, at least in part, by redox‐regulated chemical reactions in cells. Alterations in pH, pO2, cellular reductants, transition metals, and UV light lead to the loss and/or gain of S‐NO bonds. Due to the redox‐sensitive nature of the modification, analysis of protein S‐nitrosylation is technically difficult, the S‐NO bond formation being easily disrupted during sample preparation. In addition, the level of S‐nitrosylated proteins in cells approaches the limit of detection of currently available technology. Despite these technical challenges, several useful methods have been developed recently to measure protein S‐nitrosylation in biologic samples, and these are described in this unit.