bluuebunny/crossref_metadata_2025 huggingface dataset:
Short-term, but not post-exposure, protection against lethal orthopoxvirus challenge after immunization with modified vaccinia virus Ankara
Authors: Staib Caroline , Suezer Yasemin , Kisling Sigrid , Kalinke Ulrich , Sutter Gerd
Published: September 2006

Safety-tested vaccinia virus (VACV) MVA serves as a candidate third-generation vaccine against smallpox. Here, MVA immunization of mice shortly before or after lethal respiratory challenge with VACV Western Reserve was investigated. Whilst post-exposure treatment failed to protect animals, immunizations on day 2 prior to challenge were fully protective. On the day of challenge, MVA inoculation may prevent death, but not onset of severe respiratory disease. After intranasal MVA application, massive influx of leukocytes (such as neutrophils, macrophages, natural killer cells and T cells) was found in the lungs of the animals, indicating the contribution of innate responses to protection. Correspondingly, in RAG-1−/− mice, MVA inoculation delayed onset of disease significantly, but did not prevent fatal infection. Thus, short-term protection required a tight interplay of both innate and adaptive antiviral immunity. These data suggest that, in addition to conventional vaccination, MVA may serve for potent emergency prophylaxis against orthopoxvirus infection.
T-cell responses to peptide fragments of the BK virus T antigen: implications for cross-reactivity of immune response to JC virus
Authors: Li Jongming , Melenhorst Jos , Hensel Nancy , Rezvani Katyoun , Sconocchia Giuseppe , Kilical Yasemin , Hou Jean , Curfman Blanche , Major Eugene , Barrett A. John
Published: September 2006

Infection with BK virus (BKV) induces both humoral and cellular immunity, but the viral antigens of T-antigen (T-ag) stimulating T-cell responses are largely unknown. To identify BKV-specific T cells in healthy individuals, peripheral blood lymphocytes were cultured with autologous dendritic cells (DCs) loaded with BKV lysate and T cells were screened for intracellular gamma interferon production after stimulation with an overlapping 15mer peptide library of the BKV T-ag. Among many immunogenic peptides identified, four T-ag peptides were identified as candidate major histocompatibility complex class I and II T-cell epitopes, restricted to human leukocyte antigen (HLA)-B*0702, -B*08, -DRB1*0301 and -DRB1*0901. Further, a candidate 9mer peptide, LPLMRKAYL, was confirmed to be restricted to HLA-B*0702 and -B*08. Because the polyomaviruses BKV, JC virus (JCV) andSimian virus 40(SV40) share extensive sequence similarity in the immunogenic proteins T-ag and VP1, it was hypothesized that, in humans, these proteins contain conserved cytotoxic T-lymphocyte (CTL) target epitopes. Four HLA-restricted conserved epitopes of BKV, JCV and SV40 were identified: HLA-B*07, -B*08 and -DRB1*0901 for T-ag and -A*0201 for VP1. T cells culturedin vitrothat were specific for one viral antigen recognized other conserved epitopes. CTLs generated from BKV T-ag and VP1 peptide were cytotoxic to DC targets pulsed with either BKV or JCV. Therefore, infection by one of the two viruses (BKV and JCV) could establish cross-immunity against the other. Although cross-cytotoxicity experiments were not performed with SV40, cross-recognition data from conserved antigen epitopes of polyomaviruses suggest strongly that cross-immunity might also exist among the three viruses.
Virulence attenuation of Dengue virus due to augmented glycosaminoglycan-binding affinity and restriction in extraneural dissemination
Authors: Lee Eva , Wright Peter J. , Davidson Andrew , Lobigs Mario
Published: September 2006

To gain insight into the role of cell surface glycosaminoglycans (GAG) in dengue virus (DEN) cell tropism and virulence, DEN-2 mouse brain-adapted vaccine candidate, neurovirulent prototype strain (NGC) and low-passage strain, PUO-218, were passaged in BHK-21 and SW13 cells to isolate variants with high affinity for GAG. Sequence comparisons of parent and passage variants revealed five GAG-binding determinants, which all cluster in a surface-exposed region in domain II of the three-dimensional structure of the DEN envelope protein. Using an infectious cDNA clone of NGC and an NGC/PUO-218 prM–E chimeric clone, it was demonstrated that the GAG-binding determinants augment the specific infectivity for BHK-21 and/or SW13 cells by 10- to 170-fold and in some cases marginally reduce that for Vero cells. This altered cell tropism was due to a greater dependence of the variants on cell surface GAG for attachment/entry, given their increased susceptibility to heparin inhibition. The effect of the GAG-binding determinants on virulence was examined in mice deficient in alpha/beta/gamma interferon responses. High GAG affinity strongly correlated with low neuroinvasiveness due to rapid virus clearance from the blood. It was speculated that this mechanism accounts for the attenuation in primates of some DEN vaccine candidates. Interestingly, the GAG-binding variants did not display marked attenuation of neurovirulence and the opposing effect of enhanced neurovirulence was associated with one determinant (Lys126) already present in mouse brain-adapted NGC. This discrepancy of attenuated neuroinvasiveness and augmented neurovirulence may be reconciled by the existence of different mechanisms of virus dissemination in the brain and in extraneural tissues.
Glycoprotein G is a virulence factor in infectious laryngotracheitis virus
Authors: Devlin J. M. , Browning G. F. , Hartley C. A. , Kirkpatrick N. C. , Mahmoudian A. , Noormohammadi A. H. , Gilkerson J. R.
Published: September 2006

Infectious laryngotracheitis virus (ILTV; Gallid herpesvirus 1) is an alphaherpesvirus that causes acute respiratory disease in chickens. The role of glycoprotein G (gG) in vitro has been investigated in a number of alphaherpesviruses, but the relevance of gG in vivo in the pathogenicity of ILTV or in other alphaherpesviruses is unknown. In this study, gG-deficient mutants of ILTV were generated and inoculated into specific-pathogen-free chickens to assess the role of gG in pathogenicity. In chickens, gG-deficient ILTV reached a similar titre to wild-type (wt) ILTV but was significantly attenuated with respect to induction of clinical signs, effect on weight gain and bird mortality. In addition, an increased tracheal mucosal thickness, reflecting increased inflammatory cell infiltration at the site of infection, was detected in birds inoculated with gG-deficient ILTV compared with birds inoculated with wt ILTV. The reinsertion of gG into gG-deficient ILTV restored the in vivo phenotype of the mutant to that of wt ILTV. Quantitative PCR analysis of the expression of the genes adjacent to gG demonstrated that they were not affected by the deletion of gG and investigations in vitro confirmed that the phenotype of gG-deficient ILTV was consistent with unaltered expression of these adjacent genes. This is the first reported study to demonstrate definitively that gG is a virulence factor in ILTV and that deletion of gG from this alphaherpesvirus genome causes marked attenuation of the virus in its natural host.
Flavivirus membrane fusion
Authors: Stiasny Karin , Heinz Franz X.
Published: September 2006

Flavivirus membrane fusion is mediated by a class II viral fusion protein, the major envelope protein E, and the fusion process is extremely fast and efficient. Understanding of the underlying mechanisms has been advanced significantly by the determination of E protein structures in their pre- and post-fusion conformations and by the elucidation of the quarternary organization of E proteins in the viral envelope. In this review, these structural data are discussed in the context of functional and biochemical analyses of the flavivirus fusion mechanism and its characteristics are compared with those of other class II- and class I-driven fusion processes.
Measles virus: cellular receptors, tropism and pathogenesis
Authors: Yanagi Yusuke , Takeda Makoto , Ohno Shinji
Published: September 2006

Measles virus(MV), a member of the genusMorbillivirusin the familyParamyxoviridae, is an enveloped virus with a non-segmented, negative-strand RNA genome. It has two envelope glycoproteins, the haemagglutinin (H) and fusion proteins, which are responsible for attachment and membrane fusion, respectively. Human signalling lymphocyte activation molecule (SLAM; also called CD150), a membrane glycoprotein of the immunoglobulin superfamily, acts as a cellular receptor for MV. SLAM is expressed on immature thymocytes, activated lymphocytes, macrophages and dendritic cells and regulates production of interleukin (IL)-4 and IL-13 by CD4+T cells, as well as production of IL-12, tumour necrosis factor alpha and nitric oxide by macrophages. The distribution of SLAM is in accord with the lymphotropism and immunosuppressive nature of MV.Canine distemper virusandRinderpest virus, other members of the genusMorbillivirus, also use canine and bovine SLAM as receptors, respectively. Laboratory-adapted MV strains may use the ubiquitously expressed CD46, a complement-regulatory molecule, as an alternative receptor through amino acid substitutions in the H protein. Furthermore, MV can infect SLAM−cells, albeit inefficiently, via the SLAM- and CD46-independent pathway, which may account for MV infection of epithelial, endothelial and neuronal cellsin vivo. MV infection, however, is not determined entirely by the H protein–receptor interaction, and other MV proteins can also contribute to its efficient growth by facilitating virus replication at post-entry steps. Identification of SLAM as the principal receptor for MV has provided us with an important clue for better understanding of MV tropism and pathogenesis.
G1Synchronization of CHO Cells by Isoleucine Deprivation
Authors: Galgano Paul J. , Schildkraut Carl L.
Published: September 2006

INTRODUCTION

This protocol provides a method for the synchronization of a monolayer culture of CHO cells in G1using isoleucine deprivation. Since CHO cells can also be adapted to grow in suspension culture, this procedure can be used to obtain larger quantities of cells. When isoleucine is replaced, the cells resume growth and begin to enter S phase ~4 hours later. This method arrests almost 100% of the CHO cells in G1, and upon reversal, leads to rapid recovery of cell growth and very high cell viability. This method has been used with other hamster cell lines, some transformed mouse cells, and some transformed human cell lines have also been used, but with varying effectiveness.
G1/S Phase Synchronization Using Mimosine Arrest
Authors: Galgano Paul J. , Schildkraut Carl L.
Published: September 2006

INTRODUCTION

This protocol uses the plant amino acid mimosine as a G1/S synchronizing agent. Cells are first treated with excess thymidine to accumulate the majority of them at G1/S; however, some cells will have stopped growth within the S phase. Thymidine is then removed to allow all the cells to proceed completely through the S phase. Mimosine is then added to arrest the cells at the G1/S border. When mimosine is removed, cells will begin to enter S phase within about 1 hour.
Freezing and Thawing Tetrahymena Cultures
Authors: Sweet Melody T. , Allis C. David
Published: September 2006

INTRODUCTION

The method for freezing Tetrahymena has undergone major modifications over the years. The protocol given below was optimized by P.J. Bruns.
Production of dsRNA for RNAi in Avian Embryos
Authors: Stoeckli Esther T.
Published: September 2006

INTRODUCTION

This protocol describes the production of double-stranded RNA (dsRNA) from fragments of cDNAs of candidate genes. The cDNA fragments must be cloned in plasmids with a flanking SP6 and T7 promoter (e.g., pSP72 or pCRII). The plasmid is linearized and sense and antisense RNAs are produced separately by in vitro transcription. After purification, the RNA strands are annealed to yield a dsRNA molecule suitable for RNAi in avian embryos.
Dissection of Spinal Cords for Analysis of Axonal Pathfinding in dsRNA-Treated Avian Embryos
Authors: Stoeckli Esther T.
Published: September 2006

INTRODUCTION

This protocol describes how to dissect spinal cords of dsRNA-treated avian embryos for analysis of commissural axon guidance. After fixation of the spinal cord with paraformaldehyde, commissural axons are stained with Fast-DiI.
Whole-Mount Preparations for Analysis of Axonal Pathfinding in dsRNA-Treated Avian Embryos
Authors: Stoeckli Esther T.
Published: September 2006

INTRODUCTION

This protocol describes a method for staining nerve fibers in whole-mount preparations of avian embryos using an antibody against the 160-kD subunit of neurofilaments. This allows the comparison of the branching pattern of motor and sensory neurons between control and experimental embryos. This protocol has been successfully applied for embryos at different stages up to about stage 33 (7 days of incubation).
Fixing Caenorhabditis elegans in Paraformaldehyde
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

Another common method for fixing worms is to use paraformaldehyde. This method provides a gentler fixation than the Bouin’s method, but often requires the use of collagenase. This method is particularly good for examining adult worms.
Preparing Early Whole-Mount Drosophila Embryos for Immunostaining
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

The choice of fixation and permeabilization methods for examining flies depends on their stage of development. Early embryos (0-17 hours or until cuticle formation) are treated with a mixture of organic solvents, formaldehyde, and alcohols, as described here. The cuticles of late-stage embryos are usually opened by sonication. Tissues from more advanced stages of development are normally dissected by hand and then fixed and stained in a standard paraformaldehyde/detergent combination
Antibody Addition to Drosophila Specimens and Detection Using Fluorochrome-Linked Reagents
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

Fluorochrome-linked reagents should be used when high resolution is needed or if two antigens need to be localized simultaneously. Because of the thickness of fly specimens, detection requires access to a confocal microscope.
Unmasking Hidden Epitopes with Proteases
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

Fixation can mask epitopes. However, it is often possible to re-expose them using a gentle incubation with proteases, which removes obstructing structures and allows antibody access, as described here. Many proteases can be used for this procedure, including very crude preparations of proteases, such as pronase. However, using a better-characterized protease, such as trypsin, allows a more controlled reaction and better comparison between experiments. Be aware that any of the antigen retrieval methods should be avoided wherever possible, because they may introduce artifactual false-positive staining. Methods for antigen retrieval are specific for each antibody/antigen combination. No attempts to establish quantitative data following these methods should be considered.
Unmasking Hidden Epitopes Using the Pressure Cooker
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

Several methods have been developed to “retrieve” antigens that have been masked by fixation. The principle behind the pressure cooker method described here is to use extended periods of heat to break some of the subcellular structures that block antibody access. This approach is appropriate for handling specimens on glass slides. The major advantages of the pressure cooker method are the ability to handle a large number of slides simultaneously, the convenience of using metal racks, and the avoidance of any hot spots that are found in the microwave. Be aware that any of the antigen retrieval methods should be avoided wherever possible, because they may introduce artifactual false-positive staining. Methods for antigen retrieval are specific for each antibody/antigen combination. No attempts to establish quantitative data following these methods should be considered.
Immunoprecipitation: Lysing Bacteria by Sonication
Authors: Harlow Ed , Lane David
Published: September 2006

INTRODUCTION

In this protocol, bacterial cells are lysed by being subjected to short, intense treatments with ultrasound, which breaks the cell walls and shears the DNA into sizes that will not affect the viscosity of the samples. Note that this method causes some denaturation of the samples. The resulting lysate is ready for preclearing.
Fragmentation of DNA by Sonication: Figure 1.
Authors: Sambrook Joseph , Russell David W.
Published: September 2006

INTRODUCTION

DNA fragmentation is often necessary prior to library construction or subcloning for DNA sequencing. This protocol describes a method for DNA fragmentation by sonication. During sonication, DNA samples are subjected to hydrodynamic shearing by exposure to brief periods of sonication. DNA that has been sonicated for excessive periods of time is extremely difficult to clone. Most sonicators will not shear DNA to a size of less than 300-500 bp, and it is tempting to continue sonication until the entire DNA population has been reduced in size. However, the yield of subclones is usually greater if sonication is stopped when the fragments of the target DNA first reach a size of ~700 bp.
Staining SDS-Polyacrylamide Gels with Silver Salts
Authors: Sambrook Joseph , Russell David W.
Published: September 2006

INTRODUCTION

A number of methods have been developed to stain polypeptides with silver salts after separation by SDS-polyacrylamide gel electrophoresis. It is somewhat more difficult to perform than staining with Coomassie Brilliant Blue R-250, but it is ~100-1000-fold more sensitive and is capable of detecting as little as 0.1-1.0 ng of polypeptide in a single band.
Chromatin Immunoprecipitation (ChIP) of Protein Complexes: Mapping of Genomic Targets of Nuclear Proteins in Cultured Cells
Authors: Breiling Achim , Orlando Valerio
Published: September 2006

INTRODUCTION

This protocol describes the use of chromatin immunoprecipitation technology (ChIP) to analyze interactions of proteins or protein complexes with DNA in vivo. In this approach, the material is fixed with formaldehyde to preserve DNA-protein and protein-protein associations, the cells are lysed, and the chromatin is cut and solubilized. The chromatin suspension is immunoprecipitated with an antibody against the protein(s) of interest, and the coimmunoprecipitated DNA fragments are analyzed. The following protocol has been established for the cultured cell line Schneider 2 (S2) from Drosophila melanogaster. If other tissue is used, certain steps of the protocol may need to be optimized; the main variation is likely to be in the cross-linking step.
Genome-Wide Analysis of Protein-Protein Interactions Using a Two-Hybrid Array: High-Density Robotic Screening Procedure: Figure 1.
Authors: Hazbun Tony R. , Miller John P.
Published: September 2006

INTRODUCTION

This protocol describes how to handle arrayed yeast using robotics.
Analysis of Protein Interactions with Immobilized Peptide Arrays Synthesized on Membrane Supports
Authors: Frank Ronald , Dübel Stefan
Published: September 2006

INTRODUCTION

The following protocol describes the synthesis of short linear peptides, or peptide pools, on modified cellulose membranes, and the detection of their protein-binding partners. Peptides are synthesized from their carboxyl termini using Fmoc-amino acid derivatives. After completion of the synthesis and cleavage of all side-chain-protecting groups, the peptide array on the membrane is incubated with the potential interaction partners to identify their target sequences.
Global tests of haemostasis in critically ill patients with severe sepsis syndrome compared to controls
Authors: Collins Peter W. , Macchiavello Luis I. , Lewis Sarah J. , Macartney Nichola J. , Saayman Anton G. , Luddington Roger , Baglin Trevor , Findlay George P.
Published: September 2006

Haemostatic changes in septic patients are complex, with both procoagulant and anticoagulant changes. Thirty‐eight patients with severe sepsis and 32 controls were investigated by coagulation screens, individual factor assays, calibrated automated thrombography (CAT), whole blood low‐dose‐tissue factor activated (LD‐TFA) Rotem and LD‐TFA waveform analysis. Thirty‐six of 38 patients had an abnormal coagulation screen. The mean levels of factors II, V (P < 0·05), VII, X, XI and XII, antithrombin and protein C (P < 0·01) was decreased in sepsis compared with controls. The mean factor VIII and fibrinogen level (P < 0·001) was increased. CAT in platelet rich and poor plasma showed a prolonged lag time (P < 0·02), decreased peak thrombin (P < 0·02) and delayed time to peak thrombin (P < 0·001) in sepsis patients, however, the endogenous thrombin potential was equivalent in sepsis and controls. In LD‐TFA Rotem, septic patients had delayed clot times (P = 0·04) but an increased maximum velocity of clot formation (P < 0·01) and area under the clot elasticity curve (P < 0·01). LD‐TFA waveform analysis showed a delayed onset time but an increased rate of clot formation (P < 0·005). In conclusion, global tests of haemostasis suggest that in this patient group, activation of haemostasis is delayed but once initiated thrombin generation and clot formation are normal or enhanced.
Positron emission tomography/computed tomography: diagnostic accuracy in lymphoma
Authors: Hernandez‐Maraver Dolores , Hernandez‐Navarro Fernando , Gomez‐Leon Nieves , Coya Juan , Rodriguez‐Vigil Beatriz , Madero Rosario , Pinilla Immaculada , Martin‐Curto Luis M.
Published: September 2006

An accurate initial staging of patients with non‐Hodgkin lymphoma (NHL) and Hodgkin lymphoma (HL) is critical for the selection of an appropriate treatment. Computed tomography (CT) remains the standard imaging technique, although it is based on anatomic criteria. Positron emission tomography (PET) with 2‐deoxy‐2‐[fluorine‐18]fluoro‐d‐glucose (FDG) provides useful functional information but requires anatomical correlation to localise lesions accurately. We have prospectively compared the accuracy of combined PET/CT with that of CT and PET alone at initial staging in lymphoma patients. Forty‐seven newly diagnosed patients were evaluated. PET/CT was superior compared with CT and PET in nodal evaluation and detection of extranodal disease. Using a staging algorithm with PET/CT resulted in the disease stage being increased in 11 of 47 patients (10 NHL and 1 HL) (McNemar test P = 0·012). Therefore, a different treatment strategy based on PET/CT findings was suggested for seven patients (14·8%). PET/CT markedly improves accuracy in the diagnostic work‐up of patients with lymphoma.
Pure red cell aplasia associated with thymoma: clinical insights from a 50‐year single‐institution experience*
Authors: Thompson Carrie A. , Steensma David P.
Published: September 2006

Acquired pure red cell aplasia (PRCA) is a rare disorder of erythropoiesis that can develop in association with a thymoma. Optimal management of this subgroup is unclear, and there have been few series reporting long‐term clinical outcomes. Here, we report features of 13 patients treated for PRCA associated with thymoma over 50 years at our institution. Surgical resection of the thymoma was insufficient for normalisation of erythropoiesis in all cases. T‐cell gene rearrangement studies did not routinely demonstrate a clonal process, and ciclosporin and anti‐thymocyte globulin were effective adjuvant treatments. However, treatment‐related morbidity was high, with frequent infectious complications.
The membrane attack pathway of complement drives pathology in passively induced experimental autoimmune myasthenia gravis in mice
Authors: Morgan B P , Chamberlain-Banoub J , Neal J W , Song W , Mizuno M , Harris C L
Published: September 2006

The human neuromuscular disease myasthenia gravis (MG) is characterized by the generation of autoantibodies reactive with nicotinic acetylcholine receptors (AChR) that cause loss of AChR from the neuromuscular end-plate with resultant failure of neuromuscular transmission. A role for complement (C) in AChR loss has been suggested based upon morphological identification of C at the end-plate in MG and from the effects of C inhibition in murine models. Here we provide further evidence implicating C, and specifically the membrane attack complex (MAC), in a mouse model of MG. Mice deficient in the C regulators Daf1 and/or Cd59a were tested in the model. Wild-type mice were resistant to disease while mice deficient in Daf1 had mild disease symptoms with evidence of C activation and AChR loss at end-plates. Cd59a-deficient mice had very mild disease with some muscle inflammation and essentially undamaged end-plates. In contrast, mice deficient in both C regulators developed a severe paralytic disease with marked muscle inflammation and loss of end-plates. Inhibition of MAC assembly abrogated clinical disease in these double-deficient mice, demonstrating conclusively that MAC formation was driving pathology in the model. These findings provoke us to suggest that current anti-C therapeutics targeting MAC assembly will be beneficial in MG patients resistant to conventional therapies.
Adult survivors of childhood acute lymphoblastic leukaemia with GH deficiency have normal self‐rated quality of life but impaired neuropsychological performance 20 years after cranial irradiation
Authors: Link Katarina , Moëll Christian , Österberg Kai , Persson Roger , Ørbaek Palle , Garwicz Stanislaw , Cavallin‐Ståhl Eva , Erfurth Eva Marie
Published: September 2006

Objective  Cranial radiotherapy (CRT) was, until recently, important for achieving long‐term survival in acute lymphoblastic leukaemia (ALL). Because survival rates have improved markedly, the long‐term complications, such as GH deficiency (GHD) and neuropsychological impairment, have become increasingly important.

Design and patients  The level of self‐reported quality of life and neuropsychological functioning was investigated in 44 adults (21 women) with a median age of 25 years who had been treated for childhood onset (CO) ALL with CRT (median 24 Gy). Comparison was made with matched population controls. A subset of patients with GHD was evaluated for neuropsychological functioning after 1 year of GH treatment.

Results  Compared to controls, the patients had significantly lower performance in neuropsychological tests. Early age at treatment had a significant negative impact on neuropsychological performance in adulthood. No relationship was found between dose of CRT, time since treatment of ALL or gender and neuropsychological performance. Compared to controls, the patients did not show a poor quality of life or a lowered availability of social interactions or social networks; however, significantly more patients were living alone or with their parents. After GH testing, the patients were all considered GH deficient or insufficient, but no relationship was observed between stimulated peak GH secretion and neuropsychological performance. Treatment with GH for 1 year in a subgroup of the patients did not improve their neuropsychological performance.

Conclusions  This study showed that adults treated with CRT for CO ALL had GHD and significantly impaired neuropsychological performance, although self‐reported quality of life was not affected. The effect of GH treatment in this patient group has to be further elucidated.
Bone mineral density in patients with growth hormone deficiency: does a gender difference exist?
Authors: Hitz Mette Friberg , Jensen Jens‐Erik Beck , Eskildsen Peter C.
Published: September 2006

Objective  The aim of the study was to clarify whether a gender difference exists with respect to bone mineral density (BMD) and bone mineral content (BMC) in adult patients with growth hormone deficiency (GHD).

Design A case–control design.

Methods Blood sampling for measurements of calcium, phosphate, creatinine, PTH, vitamin D, IGF‐1, markers of bone formation and bone resorption, and dual energy X‐ray absorptiometry (DEXA), to determine BMD and BMC of the lumbar spine, hip, distal arm and total body, were performed in 34 patients with GHD (19 females) and 34 sex‐, age‐ and weight‐matched healthy control subjects. The patients were well substituted on all pituitary axes, apart from GH.

Results GH‐deficient males had significantly lower BMD in the lumbar spine (P = 0·02), hip (P = 0·01) and total body (P = 0·003) than healthy males while GH‐deficient females compared to healthy females had identical BMD values at all regions. This gender difference was even more obvious when BMD values were expressed asZ‐scores or as three‐dimensional BMD of the total body. The bone formation and bone resorption markers, as well as calcium and vitamin D, were all at the same levels in GH‐deficient and healthy males, indicating identical bone turnover. The GH‐deficient females, however, had significantly lower levels of bone markers compared to healthy females, indicating a reduced bone turnover. Oestrogen substitution of the GH‐deficient females could explain this difference.

Conclusions Compared to healthy control subjects GH‐deficient males had, in contrast to GH‐deficient females, significantly reduced BMD and BMC. This obvious gender difference seems to be caused by the oestrogen substitution given to the females, compensating for the lack of GH, an effect testosterone does not seem to possess.
Expression of activin and inhibin subunits, receptors and binding proteins in Human adrenocortical neoplasms
Authors: Hofland J. , Timmerman M. A. , De Herder W. W. , Van Schaik R. H. N. , De Krijger R. R. , De Jong F. H.
Published: September 2006

Objective  The growth and differentiation factors activin and inhibin can affect tumour formation and steroid production in the adrenal cortex. These factors bind to type I (Alk‐4), type II (ActRIIA, ActRIIB) and type III (betaglycan) receptors or to the activin‐binding protein follistatin. Expression of these activin‐related mRNAs was measured in different types of adrenocortical tissues and tumours to study the relationship with tumorigenesis.

Design  Quantitative expression of activin‐related mRNAs was investigated in patient adrenocortical samples.

Patients  Twenty‐eight human adrenocortical samples from normal and hyperplastic adrenals and from adrenocortical adenomas and carcinomas were collected after surgery for study purposes.

Measurements  Using quantitative reverse transcription polymerase chain reaction (RT‐PCR), we investigated the expression of inhibin α‐, βA‐ and βB‐subunits, follistatin, betaglycan, ActRIIA, ActRIIB and Alk‐4 in the adrenocortical tissues. The expression of cytochrome P450c17 (CYP17) mRNA was also measured to investigate its association with inhibin and activin subunit expression.

Results  All genes studied were expressed in all tissues, with the exception of the inhibin α‐subunit in one hyperplastic adrenal and three adrenocortical carcinomas. Expression of inhibin βA‐subunit, follistatin, betaglycan, ActRIIA, ActRIIB and CYP17 differed between nontumorous adrenals and carcinomas.

Conclusions  These differences, together with correlation analysis, indicate parallel regulation of the expression of CYP17, the inhibin α‐subunit, ActRIIA, ActRIIB, betaglycan and follistatin. We conclude that the expression of activin and inhibin subunits, receptors and binding proteins is affected by tumour formation in the adrenal gland and may play a role in tumorigenesis.
Cutaneous squamous cell carcinoma metastatic to chronic lymphocytic leukaemia: diagnostic potential of fine needle aspiration cytology
Authors: Fležar M. Strojan , Prevodnik V. Kloboves , Kirbiš I. Srebotnik , Strojan P.
Published: September 2006

Objective:  The phenomenon of cancer‐to‐cancer metastasis of cutaneous squamous cell carcinoma (SCC) and chronic lymphocytic leukaemia (CLL)/small lymphocytic lymphoma (SLL) is a rare event and only occasionally documented in the medical literature.

Methods:  Two patients with SCC of the skin that were previously treated for CLL are presented. Both had palpable lymph nodes in the neck and fine needle aspiration cytology (FNAC) was performed to evaluate the pathological process. In addition, the literature on cutaneous SCC metastatic to CLL/SLL with special emphasis on the role of FNAC in this specific clinical situation was reviewed.

Results:  On examination of the FNAC smear, cancer‐to‐cancer metastasis of cutaneous SCC to lymph node replaced by CLL was found. In one of the patients, light microscopy examination of the smear was complemented by immunostaining of atypical cells with cytokeratin antibodies and immunophenotyping of lymphoid cells by flow cytometry. In addition to our two patients, nine cases of cutaneous SCC metastatic to CLL/SLL have been reported in the literature, and in only one was the diagnosis made by FNAC.

Conclusion:  FNAC supported by ancillary immunocytological techniques could also be used in diagnosis of specific clinical situations such as cancer‐to‐cancer metastasis of the tandem of SCC–CLL/SLL.
Comparative cytomorphology of skin, lymph node, liver and bone marrow in patients with lepromatous leprosy1
Authors: Singh N. , Bhatia A. , Lakra A. , Arora V. K. , Bhattacharya S. N.
Published: September 2006

Objective:  The aim of this study was to compare the cytological changes in skin, lymph nodes, liver and bone marrow in patients with lepromatous leprosy.

Methods:  Skin lesion, lymph node, liver and bone marrow aspirates were analysed. May–Grunwald–Giemsa (MGG) and Ziehl–Neelsen (Z–N) stains were employed. Comparative cytomorphology was studied.

Results:  Twenty patients with lepromatous leprosy were studied. Lepra cells (LC) predominated in the skin aspirates of 12 patients with lepromatous leprosy (LL), lymphocytes accompanied LC in eight patients with borderline‐lepromatous (BL) leprosy. Three patients of LL leprosy and two of BL leprosy in type 2 reaction additionally had numerous neutrophils. Two patterns of lymph node aspirates were seen: partial replacement with few LC in a reactive lymphoid background (10), complete replacement with either only LC or LC in a background of degenerating neutrophils (10), the latter a feature of type 2 reaction. Liver aspiration was performed in seven patients and of bone marrow in eight patients. Occasional LC were present in five liver‐aspirated patients, steatosis and Kupffer cell hyperplasia in four patients, and myelopoiesis in two patients. Bone marrow smears invariably had occasional LC and a relative increase in mature plasma cells; sea‐blue histiocytes were seen in six patients.

Conclusion:  Lepra cells predominated in skin and lymph node aspirates with complete replacement. In comparison, liver, bone marrow and lymph node aspirates with partial replacement were dominated by a preponderance of cells native to these organs with only few or occasional LC.
Role of ancillary techniques in diagnosing and subclassifying non‐Hodgkin's lymphomas on fine needle aspiration cytology
Authors: Dey P.
Published: September 2006

Non‐Hodgkin's lymphomas (NHL) are tumours of the lymphoid cells. During the process of development of lymphoid cells, neoplasia may evolve at any point. Neoplastic cells usually carry the imprint of cell of origin at the stage of origin. Various types of NHL may have similar morphology with wide variation in origin, immunophenotype and other biological features. Different ancillary laboratory techniques may help to overcome the limitations of morphology in this aspect. The commonly used ancillary techniques in lymphomas are immunocytochemistry (IC), flow cytometry, Southern blot (SB) technique, polymerase chain reaction (PCR) and fluorescent in situ hybridization (FISH). In addition, laser scanning cytometry (LSC) and DNA microarray technologies are in the research phase. Various laboratory techniques are used for immunophenotyping, demonstration of monoclonality, identification of chromosomal translocation, assessment of cell kinetics and expression of mRNA in the tumour cells. Flow cytometry helps in rapid immunophenotying of NHL and it has an added advantage over IC in recognizing the co‐expression of CD markers. Fine needle aspiration cytology (FNAC) combined with flow immunophenotyping may help us to diagnose and subclassify certain NHLs, such as follicular lymphoma and mantle cell lymphoma, which were previously recognized as pure morphological entities. Loss of morphology is one of the important limitations of flow cytometry. LSC can overcome this limitation by studying morphology along with the immunophenotyping pattern of individual cells. Chromosomal changes in NHL can be identified by SB, PCR and FISH. Molecular diagnosis of NHL helps in diagnosis, subclassification, prognostic assessment and even in planning of therapy. DNA microarray is a relatively newer and promising technology. It gives information about the expression of several thousands of genes in a tumour in a single experiment. In the near future, FNAC combined with ancillary techniques may play a major role in diagnosis, subclassification and management of lymphomas.
Ultrasound‐guided fine needle aspiration cytology in the diagnosis and management of thyroid nodules
Authors: Cai X. J. , Valiyaparambath N. , Nixon P. , Waghorn A. , Giles T. , Helliwell T.
Published: September 2006

Objective:  To examine the accuracy of fine needle aspiration cytology (FNAC) in the diagnosis of thyroid nodules and compare the inadequacy rates for ultrasound‐guided and freehand FNAC.

Methods:  A retrospective study of 434 patients with thyroid nodules who underwent diagnostic FNAC over a 2‐year period. Cytological diagnoses have been compared with the histological assessment of resection specimens in 69 cases.

Results:  The inadequacy rate was significantly lower from ultrasound guided FNAC (24/373 cases, 6.4%) than from freehand FNAC (8/61 cases, 13.1%) (P = 0.043). Seventy‐six percentage of patients had a non‐neoplastic cytological diagnosis and, after multidisciplinary review, the patients were reassured and assigned to clinical follow‐up. Sixty‐seven patients had a resection for cytological appearances consistent with non‐neoplastic disease (n = 34), suspicious of follicular neoplasia (n = 23), or suspicious of malignancy (n = 10), and two patients had resections following inadequate cytology with ultrasound appearances suspicious of a neoplasm. The overall accuracy of FNAC analysis for malignancy was 97.0%, with sensitivity 83.3%, specificity 98.0%, positive predictive value 71.4% and negative predictive value 98.4%. The overall accuracy of FNAC analysis for the prediction of neoplasia was 97.5%, with sensitivity 80.5%, specificity 97.8%, positive predictive value 89.2% and negative predictive value 95.9%. Difficulties in cytological diagnosis were associated with lymphoid infiltrates and with degenerative changes in follicular adenomas.

Conclusion:  Ultrasound‐guided FNAC has a significantly lower yield of inadequate aspirates than palpable FNAC. The ability of FNAC to predict neoplasia in 89% patients and to exclude neoplasia in 95.9% patients makes an important contribution to the multidisciplinary assessment of patients.
Fine needle aspiration cytology: a survey of current European practice
Authors: Kocjan G. , Feichter G. , Hagmar B. , Kapila K. , Kardum‐Skelin I. , Kloboves V. , Kobayashi T. K. , Koutselini H. , Majak B. , Schenck U. , Schmitt F. , Tani E. , Totch M. , Onal B. , Vass L. , Vielh P. , Weynand B. , Herbert A.
Published: September 2006

Fine needle aspiration cytology (FNAC) is practised widely throughout Europe. The majority of countries have dedicated cytopathologists as well as histopathologists practicing cytology. Despite this, FNAC is performed mostly by clinicians and radiologists except in the larger centres with dedicated staff with a special interest in cytopathology. The advent of One‐Stop diagnostic services and image‐guided procedures are prompting further development of FNAC clinics where cytopathologists take their own samples, issue reports in the same clinical session and take extra material for ancillary tests to complete the diagnosis. The volume of FNAC work varies accordingly; in dedicated centres FNAC represents up to 80% of the workload whilst, in the majority of countries, it represents one quarter or less. Hence, the rate of inadequate FNAC varies widely, depending on the local sampling policies and the organ, but does not exceed 25% in any of the countries. The most sampled organs are breast and thyroid, followed by lymph nodes. Most countries have dedicated training in cytopathology for pathology trainees, the duration varying between 6 months and 2 years of the total training time. This discussion, focusing on European practices, highlights the heterogeneity of FNAC activity but also its success in many centres where it is practiced to a high standard, particularly in breast, thyroid and lymph node pathology. The relatively high rate of inadequate material in some centres reflects local policies and calls for greater uniformity of FNAC practice, particularly specimen sampling. To achieve this, the future direction should concentrate on specialist training, to include performing as well as interpreting FNAC, as part of the curriculum. Current emphasis on web‐based training may not provide first hand experience of the FNAC procedure and should be supplemented by attending FNAC clinics and developing the technique to its full potential.
One year follow‐up of overweight and obese hypertensive adults following intensive lifestyle therapy
Authors: Jehn M. L. , Patt M. R. , Appel L. J. , Miller E. R.
Published: September 2006

Objective  To examine the long‐term effect on weight maintenance and dietary habits of participants in a clinical trial for weight loss.

Setting  Community‐based residents living in Maryland.

Participants  Forty‐four hypertensive, overweight adults who participated in a randomized clinical trial of weight loss. Participants were randomized to an intensive ‘lifestyle’ intervention or a ‘monitoring’ group.

Main outcome measures  Weight, self‐reported current intake of fat and fruit/fibre and self‐reported barriers to maintain weight loss were assessed 1 year after the completion of the Diet, Exercise and Weight‐loss Intervention Trial (DEW‐IT) trial.

Analysis  t‐tests were used to compare groups for differences in continuous variables and chi‐square tests were used to compare groups for categorical variables.

Results  Fourty‐two of the 44 DEW‐IT subjects participated in the follow‐up study. Overall, 55% (12/19) of the lifestyle intervention group remained at or below their baseline weight at 1 year, compared with 48% (11/23) of the monitoring group (P = 0.32). However, during that year, 95% (18/19) of the lifestyle intervention group and 52% (12/23) of the monitoring group gained weight from the end of the study. Both groups reported similar intake of fruits/vegetables (servings day−1), dietary fibre (g day−1) and fat (g day−1).

Conclusions and implications  The majority of participants who lost weight during the trial regained weight during the course of 1 year. A successful intensive 2‐month programme of lifestyle modification (DEW‐IT) was ineffective for long‐term maintenance of weight loss.
The impact of percutaneous endoscopic gastrostomy feeding upon daily life in adults
Authors: Brotherton A. , Abbott J. , Aggett P.
Published: September 2006

Background  The provision of home enteral tube feeding in adults has increased in the UK. This study explored how percutaneous endoscopic gastrostomy (PEG) feeding impacts on daily lives of adult patients, from the patients’ and carers’ perspectives.

Methods  A semi‐structured interview approach was developed to obtain participants’ views of the impact of living with a PEG. A cross‐sectional qualitative purposive sampling design was employed. Thirty‐four semi‐structured interviews were conducted (15 adult patients and 19 carers) and data were analysed descriptively and thematically.

Results  Difficulties arising from PEG feeding included vomiting, diarrhoea, infection of the PEG site and leakage. The key issues that emerged included relief of pressure to consume an oral diet, disturbed sleep, restricted ability to go out, restricted choice of clothing, difficulties finding a place to feed, missing being able to eat and drink, social occasions, negative attitudes of others towards feeding and the burden placed on family members.

Conclusions  The key themes that emerged from participants were diverse and highlight a need for increased social support for both patients and their carers, planned on an individual basis.
Acculturation and education level in relation to quality of the diet: a study of Surinamese South Asian and Afro‐Caribbean residents of the Netherlands
Authors: Nicolaou M. , Van Dam R. M. , Stronks K.
Published: September 2006

Background  To consider the changes in overall diet quality following migration we examined the associations of acculturation variables and education level with diet in Surinamese South Asian and Surinamese Afro‐Caribbean origin on the one hand, and ethnic Dutch residents of the Netherlands on the other. Surinam is a former Dutch colony in South America.

Methods  We randomly selected men and women aged 35–60 years: ethnic Dutch, n = 552; South Asian, n = 306; Afro‐Caribbean, n = 660. Intakes of fruit, vegetables, red meat, fish, vegetable oils, breakfast and salt were measured using a short questionnaire that formed the basis for a ‘diet quality indicator’ score. Highest education was measured and acculturation of the Surinamese groups was assessed by age at migration, number of resident years and a scale measure of social contacts with ethnic Dutch.

Results  Compared with ethnic Dutch, both Surinamese groups scored higher on overall diet quality (P ≤ 0.001) but some aspects of diet (breakfast and salt use) were less prudent. Education was positively associated with diet quality in ethnic Dutch (P ≤ 0.01), but not consistently so in Surinamese. Associations with social contact with ethnic Dutch varied for different quality aspects of the diet. Residence duration (mean = 22 years) and age at migration (mean = 21 years) were not associated with diet.

Conclusions  A greater degree of acculturation does not necessarily lead to a less healthful diet in migrants. In addition, the association of education level with diet may differ for migrant groups.

The diet of migrants differ from host populations, suggesting that migrant groups should be considered in the development of nutrition health promotion activities.