bluuebunny/crossref_metadata_2025 huggingface dataset:
Iron fortification of rice seeds through activation of the
nicotianamine synthase
gene
Authors: Lee Sichul , Jeon Un Sil , Lee Seung Jin , Kim Yoon-Keun , Persson Daniel Pergament , Husted Søren , Schjørring Jan K. , Kakei Yusuke , Masuda Hiroshi , Nishizawa Naoko K. , An Gynheung
Published: December 2009

The most widespread dietary problem in the world is mineral deficiency. We used the nicotianamine synthase (
NAS
) gene to increase mineral contents in rice grains. Nicotianamine (NA) is a chelator of metals and a key component of metal homeostasis. We isolated activation-tagged mutant lines in which expression of a rice
NAS
gene,
OsNAS3
, was increased by introducing 35S enhancer elements. Shoots and roots of the
OsNAS3
activation-tagged plants (
OsNAS3-D1
) accumulated more Fe and Zn. Seeds from our
OsNAS3-D1
plants grown on a paddy field contained elevated amounts of Fe (2.9-fold), Zn (2.2-fold), and Cu (1.7-fold). The NA level was increased 9.6-fold in
OsNAS3-D1
seeds. Analysis by size exclusion chromatography coupled with inductively coupled plasma mass spectroscopy showed that WT and
OsNAS3-D1
seeds contained equal amounts of Fe bound to IP6, whereas
OsNAS3-D1
had 7-fold more Fe bound to a low molecular mass, which was likely NA. Furthermore, this activation led to increased tolerance to Fe and Zn deficiencies and to excess metal (Zn, Cu, and Ni) toxicities. In contrast, disruption of
OsNAS3
caused an opposite phenotype. To test the bioavailability of Fe, we fed anemic mice with either engineered or WT seeds for 4 weeks and measured their concentrations of hemoglobin and hematocrit. Mice fed with engineered seeds recovered to normal levels of hemoglobin and hematocrit within 2 weeks, whereas those that ate WT seeds remained anemic. Our results suggest that an increase in bioavailable mineral content in rice grains can be achieved by enhancing
NAS
expression.
Housing growth in and near United States protected areas limits their conservation value
Authors: Radeloff Volker C. , Stewart Susan I. , Hawbaker Todd J. , Gimmi Urs , Pidgeon Anna M. , Flather Curtis H. , Hammer Roger B. , Helmers David P.
Published: December 2009

Protected areas are crucial for biodiversity conservation because they provide safe havens for species threatened by land-use change and resulting habitat loss. However, protected areas are only effective when they stop habitat loss within their boundaries, and are connected via corridors to other wild areas. The effectiveness of protected areas is threatened by development; however, the extent of this threat is unknown. We compiled spatially-detailed housing growth data from 1940 to 2030, and quantified growth for each wilderness area, national park, and national forest in the conterminous United States. Our findings show that housing development in the United States may severely limit the ability of protected areas to function as a modern “Noah’s Ark.” Between 1940 and 2000, 28 million housing units were built within 50 km of protected areas, and 940,000 were built within national forests. Housing growth rates during the 1990s within 1 km of protected areas (20% per decade) outpaced the national average (13%). If long-term trends continue, another 17 million housing units will be built within 50 km of protected areas by 2030 (1 million within 1 km), greatly diminishing their conservation value. US protected areas are increasingly isolated, housing development in their surroundings is decreasing their effective size, and national forests are even threatened by habitat loss within their administrative boundaries. Protected areas in the United States are thus threatened similarly to those in developing countries. However, housing growth poses the main threat to protected areas in the United States whereas deforestation is the main threat in developing countries.
Control of myosin-I force sensing by alternative splicing
Authors: Laakso Joseph M. , Lewis John H. , Shuman Henry , Ostap E. Michael
Published: December 2009

Myosin-Is are molecular motors that link cellular membranes to the actin cytoskeleton, where they play roles in mechano-signal transduction and membrane trafficking. Some myosin-Is are proposed to act as force sensors, dynamically modulating their motile properties in response to changes in tension. In this study, we examined force sensing by the widely expressed myosin-I isoform, myo1b, which is alternatively spliced in its light chain binding domain (LCBD), yielding proteins with lever arms of different lengths. We found the actin-detachment kinetics of the splice isoforms to be extraordinarily tension-sensitive, with the magnitude of tension sensitivity to be related to LCBD splicing. Thus, in addition to regulating step-size, motility rates, and myosin activation, the LCBD is a key regulator of force sensing. We also found that myo1b is substantially more tension-sensitive than other myosins with similar length lever arms, indicating that different myosins have different tension-sensitive transitions.
VEGFR1–mediated pericyte ablation links VEGF and PlGF to cancer-associated retinopathy
Authors: Cao Renhai , Xue Yuan , Hedlund Eva-Maria , Zhong Zhaodong , Tritsaris Katerina , Tondelli Barbara , Lucchini Franco , Zhu Zhenping , Dissing Steen , Cao Yihai
Published: December 2009

VEGF coordinates complex regulation of cellular regeneration and interactions between endothelial and perivascular cells; dysfunction of the VEGF signaling system leads to retinopathy. Here, we show that systemic delivery of VEGF and placental growth factor (PlGF) by protein implantation, tumors, and adenoviral vectors ablates pericytes from the mature retinal vasculature through the VEGF receptor 1 (VEGFR1)-mediated signaling pathway, leading to increased vascular leakage. In contrast, we demonstrate VEGF receptor 2 (VEGFR2) is primarily expressed in nonvascular photoreceptors and ganglion cells. Moreover, blockade of VEGFR1 but not VEGFR2 significantly restores pericyte saturation in mature retinal vessels. Our findings link VEGF and PlGF to cancer-associated retinopathy, reveal the molecular mechanisms of VEGFR1 ligand-mediated retinopathy, and define VEGFR1 as an important target of antiangiogenic therapy for treatment of retinopathy.
EBNA1 regulates cellular gene expression by binding cellular promoters
Authors: Canaan Allon , Haviv Izhak , Urban Alexander E. , Schulz Vincent P. , Hartman Steve , Zhang Zhengdong , Palejev Dean , Deisseroth Albert B. , Lacy Jill , Snyder Michael , Gerstein Mark , Weissman Sherman M.
Published: December 2009

Epstein–Barr virus (EBV) is associated with several types of lymphomas and epithelial tumors including Burkitt's lymphoma (BL), HIV-associated lymphoma, posttransplant lymphoproliferative disorder, and nasopharyngeal carcinoma. EBV nuclear antigen 1 (EBNA1) is expressed in all EBV associated tumors and is required for latency and transformation. EBNA1 initiates latent viral replication in B cells, maintains the viral genome copy number, and regulates transcription of other EBV-encoded latent genes. These activities are mediated through the ability of EBNA1 to bind viral-DNA. To further elucidate the role of EBNA1 in the host cell, we have examined the effect of EBNA1 on cellular gene expression by microarray analysis using the B cell BJAB and the epithelial 293 cell lines transfected with EBNA1. Analysis of the data revealed distinct profiles of cellular gene changes in BJAB and 293 cell lines. Subsequently, chromatin immune-precipitation revealed a direct binding of EBNA1 to cellular promoters. We have correlated EBNA1 bound promoters with changes in gene expression. Sequence analysis of the 100 promoters most enriched revealed a DNA motif that differs from the EBNA1 binding site in the EBV genome.
AP-1 homolog BZLF1 of Epstein–Barr virus has two essential functions dependent on the epigenetic state of the viral genome
Authors: Kalla Markus , Schmeinck Anne , Bergbauer Martin , Pich Dagmar , Hammerschmidt Wolfgang
Published: December 2009

EBV, a member of the herpes virus family, is a paradigm for human tumor viruses and a model of viral latency amenable for study in vitro. It induces resting human B lymphocytes to proliferate indefinitely in vitro and initially establishes a strictly latent infection in these cells.
BZLF1
, related to the cellular activating protein 1 (AP-1) family of transcription factors, is the viral master gene essential and sufficient to mediate the switch to induce the EBV lytic phase in latently infected B cells. Enigmatically, after infection
BZLF1
is expressed very early in the majority of primary B cells, but its early expression fails to induce the EBV lytic phase. We show that the early expression of
BZLF1
has a critical role in driving the proliferation of quiescent naïve and memory B cells but not of activated germinal center B cells.
BZLF1’s
initial failure to induce the EBV lytic phase relies on the viral DNA at first being unmethylated. We have found that the eventual and inevitable methylation of viral DNA is a prerequisite for productive infection in stably, latently infected B cells which then yield progeny virus lacking cytosine-phosphatidyl-guanosine (CpG) methylation. This progeny virus then can repeat EBV’s epigenetically regulated, biphasic life cycle. Our data indicate that the viral BZLF1 protein is crucial both to establish latency and to escape from it. Our data also indicate that EBV has evolved to appropriate its host’s mode of methylating DNA for its own epigenetic regulation.
IL-21 imposes a type II EBV gene expression on type III and type I B cells by the repression of C- and activation of LMP-1-promoter
Authors: Kis Loránd L. , Salamon Daniel , Persson Emma K. , Nagy Noémi , Scheeren Ferenc A. , Spits Hergen , Klein George , Klein Eva
Published: December 2009

Epstein–Barr virus (EBV) is associated with a variety of human tumors. Although the EBV-infected normal B cells in vitro and the EBV-carrying B cell lymphomas in immunodeficient patients express the full set of latent proteins (type III latency), the majority of EBV-associated malignancies express the restricted type I (EBNA-1 only) or type II (EBNA-1 and LMPs) viral program. The mechanisms responsible for these different latent viral gene expression patterns are only partially known. IL-21 is a potent B cell activator and plasma cell differentiation-inducer cytokine produced by CD4
+
T cells. We studied its effect on EBV-carrying B cells. In type I Burkitt lymphoma (BL) cell lines and in the conditional lymphoblastoid cell line (LCL) ER/EB2-5, IL-21 potently activated STAT3 and induced the expression of LMP-1, but not EBNA-2. The IL-21-treated type I Jijoye M13 BL line ceased to proliferate, and this was paralleled by the induction of IRF4 and the down-regulation of BCL6 expression. In the type III LCLs and BL lines, IL-21 repressed the C-promoter-derived and LMP-2A mRNAs, whereas it up-regulated the expression of LMP-1 mRNAs. The IL-21-treated type III cells underwent plasma cell differentiation with the induction of Blimp-1, and high levels of Ig and Oct-2. IL-21 might be involved in the EBNA-2-independent expression of LMP-1 in EBV-carrying type II cells. In light of the fact that IL-21 is already in clinical trials for the treatment of multiple malignancies, the in vivo modulation of EBV gene expression by IL-21 might have therapeutic benefits for the EBV-carrying malignancies.
Loss of autophagy in erythroid cells leads to defective removal of mitochondria and severe anemia in vivo
Authors: Mortensen M. , Ferguson D.J.P. , Edelmann M. , Kessler B. , Morten K.J. , Komatsu M. , Simon A.K.
Published: December 2009

Timely elimination of damaged mitochondria is essential to protect cells from the potential harm of disordered mitochondrial metabolism and release of proapoptotic proteins. In mammalian red blood cells, the expulsion of the nucleus followed by the removal of other organelles, such as mitochondria, are necessary differentiation steps. Mitochondrial sequestration by autophagosomes, followed by delivery to the lysosomal compartment for degradation (mitophagy), is a major mechanism of mitochondrial turnover. Here we show that mice lacking the essential autophagy gene
Atg7
in the hematopoietic system develop severe anemia.

Atg7
−/−

erythrocytes accumulate damaged mitochondria with altered membrane potential leading to cell death. We find that mitochondrial loss is initiated in the bone marrow at the Ter119
+
/CD71
High
stage. Proteomic analysis of erythrocyte ghosts suggests that in the absence of autophagy other cellular degradation mechanisms are induced. Importantly, neither the removal of endoplasmic reticulum nor ribosomes is affected by the lack of
Atg7
. Atg7 deficiency also led to severe lymphopenia as a result of mitochondrial damage followed by apoptosis in mature T lymphocytes. Ex vivo short-lived hematopoietic cells such as monocytes and dendritic cells were not affected by the loss of Atg7. In summary, we show that the selective removal of mitochondria by autophagy, but not other organelles, during erythropoeisis is essential and that this is a necessary developmental step in erythroid cells.
Phospholipase C-γ1 is involved in signaling the activation by high NaCl of the osmoprotective transcription factor TonEBP/OREBP
Authors: Irarrazabal Carlos E. , Gallazzini Morgan , Schnetz Michael P. , Kunin Margarita , Simons Brigitte L. , Williams Chester K. , Burg Maurice B. , Ferraris Joan D.
Published: December 2009

High NaCl elevates activity of the osmoprotective transcription factor TonEBP/OREBP by increasing its phosphorylation, transactivating activity, and localization to the nucleus. We investigated the possible role in this activation of phospholipase C-γ1 (PLC-γ1), which has a predicted binding site at TonEBP/OREBP-phospho-Y143. We find the following. (i) Activation of TonEBP/OREBP transcriptional activity by high NaCl is reduced in PLC-γ1 null cells and in HEK293 cells in which PLC-γ1 is knocked down by a specific siRNA. (ii) High NaCl increases phosphorylation of TonEBP/OREBP at Y143. (iii) Wild-type PLC-γ1 coimmunoprecipitates with wild-type TonEBP/OREBP but not TonEBP/OREBP-Y143A, and the coimmunoprecipitation is increased by high NaCl. (iv) PLC-γ1 is part of the protein complex that associates with TonEBP/OREBP at its DNA binding site. (v) Knockdown of PLC-γ1 or overexpression of a PLC-γ1-SH3 deletion mutant reduces high NaCl-dependent TonEBP/OREBP transactivating activity. (vi) Nuclear localization of PLC-γ1 is increased by high NaCl. (vii) High NaCl-induced nuclear localization of TonEBP/OREBP is reduced if cells lack PLC-γ1, if PLC-γ1 mutated in its SH2C domain is overexpressed, or if Y143 in TonEBP/OREBP is mutated to alanine. (viii) Expression of recombinant PLC-γ1 restores nuclear localization of wild-type TonEBP/OREBP in PLC-γ1 null cells but not of TonEBP/OREBP-Y143A. (ix) The PLC-γ1 phospholipase inhibitor U72133 inhibits nuclear localization of TonEBP/OREBP but not the increase of its transactivating activity. We conclude that, when NaCl is elevated, TonEBP/OREBP becomes phosphorylated at Y143, resulting in binding of PLC-γ1 to that site, which contributes to TonEBP/OREBP transcriptional activity, transactivating activity, and nuclear localization.
Genomic scale analysis of racial impact on response to IFN-α
Authors: Pos Zoltan , Selleri Silvia , Spivey Tara L. , Wang Jeanne K. , Liu Hui , Worschech Andrea , Sabatino Marianna , Monaco Alessandro , Leitman Susan F. , Falus Andras , Wang Ena , Alter Harvey J. , Marincola Francesco M.
Published: December 2009

Limited responsiveness to IFN-α in hepatitis C virus (HCV)-infected African-Americans compared to European Americans (AAs vs. EAs) hinders the management of HCV. Here, we studied healthy non-HCV-infected AA and EA subjects to test whether immune cell response to IFN-α is determined directly by race. We compared baseline and IFN-α-induced signal transducer and activator of transcription (STAT)-1, STAT-2, STAT-3, STAT-4, and STAT-5 protein and phosphorylation levels in purified T cells, global transcription, and a genomewide single-nucleotide polymorphism (SNP) profile of healthy AA and EA blood donors. In contrast to HCV-infected individuals, healthy AAs displayed no evidence of reduced STAT activation or IFN-α-stimulated gene expression compared to EAs. Although >200 genes reacted to IFN-α treatment, race had no impact on any of them. The only gene differentially expressed by the two races (NUDT3,
P
< 10
−7
) was not affected by IFN-α and bears no known relationship to IFN-α signaling or HCV pathogenesis. Genomewide analysis confirmed the self-proclaimed racial attribution of most donors, and numerous race-associated SNPs were identified within loci involved in IFN-α signaling, although they clearly did not affect responsiveness in the absence of HCV. We conclude that racial differences observed in HCV-infected patients in the responsiveness to IFN-α are unrelated to inherent racial differences in IFN-α signaling and more likely due to polymorphisms affecting the hosts’ response to HCV, which in turn may lead to a distinct disease pathophysiology responsible for altered IFN signaling and treatment response.
The RNF8/RNF168 ubiquitin ligase cascade facilitates class switch recombination
Authors: Ramachandran Shaliny , Chahwan Richard , Nepal Rajeev M. , Frieder Darina , Panier Stephanie , Roa Sergio , Zaheen Ahmad , Durocher Daniel , Scharff Matthew D. , Martin Alberto
Published: December 2009

An effective immune response requires B cells to produce several classes of antibodies through the process of class switch recombination (CSR). Activation-induced cytidine deaminase initiates CSR by deaminating deoxycytidines at switch regions within the Ig locus. This activity leads to double-stranded DNA break formation at the donor and recipient switch regions that are subsequently synapsed and ligated in a 53BP1-dependent process that remains poorly understood. The DNA damage response E3 ubiquitin ligases RNF8 and RNF168 were recently shown to facilitate recruitment of 53BP1 to sites of DNA damage. Here we show that the ubiquitination pathway mediated by RNF8 and RNF168 plays an integral part in CSR. Using the CH12F3-2 mouse B cell line that undergoes CSR to IgA at high rates, we demonstrate that knockdown of RNF8, RNF168, and 53BP1 leads to a significant decrease in CSR. We also show that 53BP1-deficient CH12F3-2 cells are protected from apoptosis mediated by the MDM2 inhibitor Nutlin-3. In contrast, deficiency in either E3 ubiquitin ligase does not protect cells from Nutlin-3–mediated apoptosis, indicating that RNF8 and RNF168 do not regulate all functions of 53BP1.
Alzheimer Aβ Peptide Induces Chromosome Mis-Segregation and Aneuploidy, Including Trisomy 21: Requirement for Tau and APP
Authors: Granic Antoneta , Padmanabhan Jaya , Norden Michelle , Potter Huntington
Published: December 2009

Both sporadic and familial Alzheimer's disease (AD) patients exhibit increased chromosome aneuploidy, particularly trisomy 21, in neurons and other cells. Significantly, trisomy 21/Down syndrome patients develop early onset AD pathology. We investigated the mechanism underlying mosaic chromosome aneuploidy in AD and report that FAD mutations in the Alzheimer Amyloid Precursor Protein gene, APP, induce chromosome mis-segregation and aneuploidy in transgenic mice and in transfected cells. Furthermore, adding synthetic Aβ peptide, the pathogenic product of APP, to cultured cells causes rapid and robust chromosome mis-segregation leading to aneuploid, including trisomy 21, daughters, which is prevented by LiCl addition or Ca2+ chelation and is replicated in tau KO cells, implicating GSK-3β, calpain, and Tau-dependent microtubule transport in the aneugenic activity of Aβ. Furthermore, APP KO cells are resistant to the aneugenic activity of Aβ, as they have been shown previously to be resistant to Aβ-induced tau phosphorylation and cell toxicity. These results indicate that Aβ-induced microtubule dysfunction leads to aneuploid neurons and may thereby contribute to the pathogenesis of AD.
Orthologs of the Class A4 Heat Shock Transcription Factor HsfA4a Confer Cadmium Tolerance in Wheat and Rice
Authors: Shim Donghwan , Hwang Jae-Ung , Lee Joohyun , Lee Sichul , Choi Yunjung , An Gynheung , Martinoia Enrico , Lee Youngsook
Published: December 2009

Cadmium (Cd) is a widespread soil pollutant; thus, the underlying molecular controls of plant Cd tolerance are of substantial interest. A screen for wheat (Triticum aestivum) genes that confer Cd tolerance to a Cd hypersensitive yeast strain identified Heat shock transcription factor A4a (HsfA4a). Ta HsfA4a is most similar to the class A4 Hsfs from monocots. The most closely related rice (Oryza sativa) homolog, Os HsfA4a, conferred Cd tolerance in yeast, as did Ta HsfA4a, but the second most closely related rice homolog, Os HsfA4d, did not. Cd tolerance was enhanced in rice plants expressing Ta HsfA4a and decreased in rice plants with knocked-down expression of Os HsfA4a. An analysis of the functional domain using chimeric proteins constructed from Ta HsfA4a and Os HsfA4d revealed that the DNA binding domain (DBD) of HsfA4a is critical for Cd tolerance, and within the DBD, Ala-31 and Leu-42 are important for Cd tolerance. Moreover, Ta HsfA4a–mediated Cd resistance in yeast requires metallothionein (MT). In the roots of wheat and rice, Cd stress caused increases in HsfA4a expression, together the MT genes. Our findings thus suggest that HsfA4a of wheat and rice confers Cd tolerance by upregulating MT gene expression in planta.
The Small Subunit of Snapdragon Geranyl Diphosphate Synthase Modifies the Chain Length Specificity of Tobacco Geranylgeranyl Diphosphate Synthase in Planta
Authors: Orlova Irina , Nagegowda Dinesh A. , Kish Christine M. , Gutensohn Michael , Maeda Hiroshi , Varbanova Marina , Fridman Eyal , Yamaguchi Shinjiro , Hanada Atsushi , Kamiya Yuji , Krichevsky Alexander , Citovsky Vitaly , Pichersky Eran , Dudareva Natalia
Published: December 2009

Geranyl diphosphate (GPP), the precursor of many monoterpene end products, is synthesized in plastids by a condensation of dimethylallyl diphosphate and isopentenyl diphosphate (IPP) in a reaction catalyzed by homodimeric or heterodimeric GPP synthase (GPPS). In the heterodimeric enzymes, a noncatalytic small subunit (GPPS.SSU) determines the product specificity of the catalytic large subunit, which may be either an active geranylgeranyl diphosphate synthase (GGPPS) or an inactive GGPPS-like protein. Here, we show that expression of snapdragon (Antirrhinum majus) GPPS.SSU in tobacco (Nicotiana tabacum) plants increased the total GPPS activity and monoterpene emission from leaves and flowers, indicating that the introduced catalytically inactive GPPS.SSU found endogenous large subunit partner(s) and formed an active snapdragon/tobacco GPPS in planta. Bimolecular fluorescence complementation and in vitro enzyme analysis of individual and hybrid proteins revealed that two of four GGPPS-like candidates from tobacco EST databases encode bona fide GGPPS that can interact with snapdragon GPPS.SSU and form a functional GPPS enzyme in plastids. The formation of chimeric GPPS in transgenic plants also resulted in leaf chlorosis, increased light sensitivity, and dwarfism due to decreased levels of chlorophylls, carotenoids, and gibberellins. In addition, these transgenic plants had reduced levels of sesquiterpene emission, suggesting that the export of isoprenoid intermediates from the plastids into the cytosol was decreased. These results provide genetic evidence that GPPS.SSU modifies the chain length specificity of phylogenetically distant GGPPS and can modulate IPP flux distribution between GPP and GGPP synthesis in planta.
Breeding requirements of Nahan’s Francolin, Francolinus nahan, in Budongo forest reserve, Uganda
Authors: Sande Eric , Dranzoa Christine , Wegge Per , Carroll John P.
Published: December 2009

The Nahan’s Francolin Francolinus nahani is a globally threatened species associated with remnant forests of the eastern equatorial lowlands of the Democratic Republic of the Congo and Uganda. We studied the species in the Budongo Forest Reserve during March 1998 to January 2000 in an attempt to document some aspects of breeding requirements. We located nests by physically searching the forest and through following radio‐tagged adults. We measured demographical and habitat parameters associated with each nest. Our data suggest that trees of large diameter at breast height with appropriate buttress formation are important for long‐time survival of Nahan’s Francolins.
Has the final countdown to wildlife extinction in Northern Central African Republic begun?
Authors: Bouché Philippe , Renaud Pierre‐Cyril , Lejeune Philippe , Vermeulen Cédric , Froment Jean‐Marc , Bangara Alfred , Fiongai Okclefort , Abdoulaye Antoine , Abakar Raymond , Fay Mike
Published: December 2009

The wildlife populations of Northern Central African Republic experienced precipitous declines during the 1970s and 1980s. While anecdotes coming out of the region indicate that the wildlife populations remain under serious threat, little is known about their status. An aerial sample count was carried out in the Northern Central African Republic at the end of the dry season in June 2005 and covered an 85,000 km2complex landscape containing national parks, hunting reserves and community hunting areas. Results show a dramatic decline of wildlife since the previous survey in 1985. In 20 years, large mammals’ numbers decreased by 65%, probably because of poaching and diseases brought by illegal cattle transhumance. Elephant (Loxodonta africana) and Buffon kob (Kobus kob) populations showed the greatest decline (over 80% each), while buffalo (Syncerus caffer), roan antelope (Hippotragus equinus) and Giant Lord’s Derby Eland (Taurotragus derbianus) populations seem stable or increasing over these last 20 years. The analysis of the wildlife population distribution by status of the different types of protected areas (national parks, hunting areas) showed that individual encounter rates of elephant and buffalo were lower in national parks than in neighbouring hunting areas, while those for roan, giraffe (Giraffa camelopardalis) and Buffon kob were higher in the national parks.
Blistering skin diseases: a bridge between dermatopathology and molecular biology
Authors: Nagy Nikoletta , McGrath John A
Published: December 2009

Nagy N & McGrath J A (2010) Histopathology56, 91–99 Blistering skin diseases: a bridge between dermatopathology and molecular biology

Although dermatopathology and molecular biology are often considered to be separate laboratory disciplines, the respective approaches are far from mutually exclusive. This is certainly the case for understanding the pathology of blistering skin diseases, both acquired and inherited. For example, in toxic epidermal necrolysis, dermatopathology in isolation may provide few clues to disease pathogenesis. There is widespread keratinocyte apoptosis and a variable infiltrate of cytotoxic T cells, but morphology alone offers little insight into what causes the epidermal destruction. In contrast, molecular biology studies have revealed several key processes that help explain the keratinocyte death, including increased expression of death receptors and their ligands on keratinocyte cell membranes as well as the presence of local or systemic immunocyte‐derived cytolytic granules. For some inherited blistering diseases, however, such as epidermolysis bullosa, the molecular pathology is complex and difficult to unravel in isolation, yet the addition of dermatopathology is helpful in focusing molecular investigations. Notably, immunolabelling of cell adhesion proteins using specific antibody probes can identify reduced or absent immunoreactivity for candidate genes/proteins. Bridging dermatopathology and molecular biology investigations facilitates a greater understanding of disease processes, improves diagnostic accuracy, and provides a basis for the development and appraisal of new treatments.
The histological assessment of cutaneous vasculitis
Authors: Carlson J Andrew
Published: December 2009

Carlson J A (2010) Histopathology56, 3–23 The histological assessment of cutaneous vasculitis

Vasculitis is defined as inflammation directed at vessels, which compromises or destroys the vessel wall leading to haemorrhagic and/or ischaemic events. Skin biopsy is the gold standard for the diagnosis of cutaneous vasculitis, whose manifestations include urticaria, infiltrative erythema, petechiae, purpura, purpuric papules, haemorrhagic vesicles and bullae, nodules, livedo racemosa, deep (punched out) ulcers and digital gangrene. These varied morphologies are a direct reflection of size of the vessels and extent of the vascular bed affected, ranging from a vasculitis affecting few superficial, small vessels in petechial eruptions to extensive pan‐dermal small vessel vasculitis in haemorrhagic bullae to muscular vessel vasculitis in lower extremity nodules with livedo racemosa. Skin biopsy, extending to subcutis and taken from the earliest, most symptomatic, reddish or purpuric lesion is crucial for obtaining a high‐yielding diagnostic sample. Based on histology, vasculitis can be classified on the size of vessels affected and the dominant immune cell mediating the inflammation (e.g. neutrophilic, granulomatous, lymphocytic, or eosinophilic). Disruption of small vessels by inflammatory cells, deposition of fibrin within the lumen and/or vessel wall coupled with nuclear debris allows for the confident recognition of small vessel, mostly neutrophilic vasculitis (also known as leukocytoclastic vasculitis). In contrast, muscular vessel vasculitis can be identified solely by infiltration of its wall by inflammatory cells. Extravasation of red blood cells (purpura) and necrosis are supportive, but not diagnostic of vasculitis as they are also seen in haemorrhagic and/or vaso‐occlusive disorders (pseudovasculitis). Vasculitic foci associated with extravascular granulomas (palisaded neutrophilic and granulomatous dermatitis), tissue eosinophilia, or tissue neutrophilia signal the risk for, or co‐existence of systemic disease. This essential histological information coupled with direct immunofluorescence and anti‐neutrophil cytoplasmic data and clinical findings enables more precise and accurate diagnosis of localized and systemic vasculitis syndromes.
Dysplastic naevi: an update
Authors: Elder David E
Published: December 2009

Elder D E (2010) Histopathology56, 112–120 Dysplastic naevi: an update

Dysplastic naevi are clinically atypical and histologically are characterized by architectural disorder and cytological atypia. Their diagnosis is reproducible if criteria and thresholds are agreed upon. They are significant only in relation to melanoma, as simulants of melanoma, as markers of individuals at increased risk of developing melanoma, and as potential and occasional actual precursors of melanoma. Morphologically and biologically, they are intermediate between common naevi and melanoma. Individuals with dysplastic naevi may have deficient DNA repair, and dysplastic naevi lesions are associated with overexpression of pheomelanin, which may lead to increased oxidative damage and increased potential for DNA damage and tumour progression.
Molecular cytogenetics of cutaneous melanocytic lesions – diagnostic, prognostic and therapeutic aspects
Authors: Blokx Willeke Am M , Van Dijk Marcory C R F , Ruiter Dirk J
Published: December 2009

Blokx W A M, van Dijk M C R F & Ruiter D J (2010) Histopathology56, 121–132 Molecular cytogenetics of cutaneous melanocytic lesions – diagnostic, prognostic and therapeutic aspects

This review intends to update current knowledge regarding molecular cytogenetics in melanocytic tumours with a focus on cutaneous melanocytic lesions. Advantages and limitations of diverse, already established methods, such as (fluorescence) in situ hybridization and mutation analysis, to detect these cytogenetic alterations in melanocytic tumours are described. In addition, the potential value of more novel techniques such as multiplex ligation‐dependent probe amplification is pointed out. This review demonstrates that at present cytogenetics has mainly increased our understanding of the pathogenesis of melanocytic tumours, with an important role for activation of the mitogen‐activated protein kinase (MAPK) signalling pathway in the initiation of melanocytic tumours. Mutations in BRAF (in common naevocellular naevi), NRAS (congenital naevi), HRAS (Spitz naevi) and GNAQ (blue naevi) can all cause MAPK activation. All these mutations seem early events in the development of melanocytic tumours, but by themselves are insufficient to cause progression towards melanoma. Additional molecular alterations are implicated in progression towards melanoma, with different genetic alterations in melanomas at different sites and with varying levels of sun exposure. This genetic heterogeneity in distinct types of naevi and melanomas can be used for the development of molecular tests for diagnostic purposes. However, at the moment only few molecular tests have become of diagnostic value and are performed in daily routine practice. This is caused by lack of large prospective studies on the diagnostic value of molecular tests including follow‐up, and by the low prevalence of certain molecular alterations. For the future we foresee an increasing role for cytogenetics in the treatment of melanoma patients with the increasing availability of targeted therapy. Potential targets for metastatic melanoma include genes involved in the MAPK pathway, such as BRAF and RAS. More recently, KIT has emerged as a potential target in melanoma patients. These targeted treatments all need careful evaluation, but might be a promising adjunct for treatment of metastatic melanoma patients, in which other therapies have not brought important survival advantages yet.
Classification of cutaneous lymphomas – an update
Authors: Kempf Werner , Sander Christian A.
Published: December 2009

Kempf W & Sander C A (2010) Histopathology56, 57–70 Classification of cutaneous lymphomas – an update

This review focuses on the evolution and conceptual aspects of classifications for cutaneous lymphomas. The World Health Organization/European Organization for Research and Treatment of Cancer (WHO/EORTC) classification and the WHO classification (4th edn, 2008) represent the first widely accepted classifications for lymphomas, in which the complete spectrum of primary cutaneous lymphomas is included. These classifications for primary cutaneous lymphomas define disease entities with distinct clinical, histological, immunophenotypic and genetic features. Final diagnosis is based on a synoptic integration of these features and implies clinicopathological correlation as a pivotal element of the diagnostic approach for primary cutaneous lymphomas. The entities, their definitions and diagnostic criteria of cutaneous lymphomas listed in the WHO/EORTC and WHO classifications are presented. Recent changes in the terminology and staging, practical implications and future perspectives are discussed.
BOB.1, CD79a and cyclin E are the most appropriate markers to discriminate classical Hodgkin’s lymphoma from primary mediastinal large B‐cell lymphoma
Authors: Hoeller Sylvia , Zihler Deborah , Zlobec Inti , Obermann Ellen C , Pileri Stefano A , Dirnhofer Stephan , Tzankov Alexandar
Published: December 2009

Hoeller S, Zihler D, Zlobec I, Obermann EC, Pileri SA, Dirnhofer S & Tzankov A (2010) Histopathology56, 217–228

BOB.1, CD79a and cyclin E are the most appropriate markers to discriminate classical Hodgkin’s lymphoma from primary mediastinal large B‐cell lymphoma

Aims:  To clarify which immunohistochemical markers could be helpful in distinguishing between classical Hodgkin’s lymphoma (cHL) and primary mediastinal B‐cell lymphoma (PMBCL) to more narrowly define ‘B‐cell lymphoma, unclassifiable, with features intermediate between diffuse large B‐cell lymphoma and cHL’.

Methods and results:  Two hundred and 83 cHLs and 51 PMBCLs were analysed on validated tissue microarray platforms with antibodies to BOB.1, CD15, CD20, CD23, CD30, CD79a, cyclin E, LMP‐1, MUM1p, p63 and Oct2. The marker cut‐off scores were calculated using receiver–operating characteristic curves. Markers with the highest positive predictive value for cHL were: CD15, cyclin E, LMP‐1 (all 100%), MUM1p (93%) and CD30 (83%). High sensitivity was achieved only by CD30 (92%) and cyclin E (79%). Nineteen percent of PMBCLs were also positive for CD30, which led to a lower specificity of CD30 as regards cHL (81%) compared with cyclin E (100%). The antibodies with the highest positive predictive value for PMBCL were: CD23 (98%), p63 (96%), BOB.1 (94%) and CD79a (90%), with high sensitivity for BOB.1 (100%), CD79a (89%) and p63 (82%).

Conclusions:  The use of at least three of the most accurate immunohistochemical markers, cyclin E, CD79a and BOB.1, may be helpful in the differential diagnosis of cHL and PMBCL.
Molecular alterations in AKT1, AKT2 and AKT3 detected in breast and prostatic cancer by FISH
Authors: Kirkegaard Tove , Witton Caroline J , Edwards Joanne , Nielsen Kirsten Vang , Jensen Linda Boye , Campbell Fiona M , Cooke Timothy G , Bartlett John M S
Published: December 2009

Kirkegaard T, Witton C J, Edwards J, Nielsen K V, Jensen L B, Campbell F M, Cooke T G & Bartlett J M S (2010) Histopathology56, 203–211

Molecular alterations in AKT1, AKT2 and AKT3 detected in breast and prostatic cancer by FISH

Aims:  The AKT family is implicated in cancer progression. There are three mammalian AKT isoforms located on chromosomes 14, 19 and 1, respectively. The aim of the study was to investigate genetic alterations of AKT in breast and prostatic cancers using fluorescence in situ hybridization (FISH).

Methods and results:  In oestrogen receptor(ER)‐positive breast carcinomas, AKT1 was deleted in five (4.8%) and amplified in one (1%) carcinoma. Deletions of AKT2 were seen in 19 (21.1%) cases. No AKT2 amplifications were identified. Ten (9.9%) AKT3 amplifications but no deletions were seen. In prostatic cancer, AKT1 was amplified in one carcinoma (2.6%). No genetic changes were observed for AKT2 and AKT3. High frequencies of aneusomy for all chromosomes were observed in breast and prostatic carcinomas.

Conclusions:  In breast cancer AKT3 amplifications and AKT1 and AKT2 deletions were seen, which, to our knowledge, have not been shown by FISH before. Although these two cohorts cannot be directly compared, only one AKT1 amplification was identified in prostatic carcinomas. This indicates differences in the genetic changes underlying development of breast and prostatic cancers. To evaluate further the role of genetic changes of AKT in breast cancer progression, a cohort of both ER+ and ER− patients should be evaluated.
Clinicopathological significance of nuclear PTEN expression in colorectal adenocarcinoma
Authors: Jang Ki‐Seok , Song Young Soo , Jang Si‐Hyong , Min Kyueng‐Whan , Na Woong , Jang Se Min , Jun Young Jin , Lee Kang Hong , Choi Dongho , Paik Seung Sam
Published: December 2009

Jang K‐S, Song Y S, Jang S‐H, Min K‐W, Na W, Jang S M, Jun Y J, Lee K H, Choi D & Paik S S (2010) Histopathology56, 229–239

Clinicopathological significance of nuclear PTEN expression in colorectal adenocarcinoma

Aims:  Tumour suppressor phosphatase and tensin homologue (PTEN) is an important negative regulator for the PIP3/Akt signalling pathway that promotes cell proliferation and inhibits apoptosis. Inactivation of PTEN by mutation, deletion and promoter hypermethylation has been demonstrated in a range of cancers. The aim was to investigate whether the loss of nuclear PTEN protein expression correlates with conventional clinicopathological parameters and patient survival.

Methods and results:  Immunohistochemistry staining for PTEN was performed on a tissue microarray of 19 samples of normal colonic mucosa, 14 adenomatous polyps, 482 adenocarcinomas and 56 metastatic lymph nodes. All 19 normal colonic mucosa samples (100%) were positive and 12 (85.7%) out of 14 adenomatous polyps were positive for PTEN. However, only 241 (50.0%) of the 482 colorectal adenocarcinomas and 26 (46.4%) of the 56 metastatic lymph nodes were positive for PTEN. Loss of PTEN expression was related to defective mismatch repair protein expression and colonic localization rather than rectal localization. On univariate survival analysis, patients with PTEN− adenocarcinoma revealed a poor overall and disease‐free survival (P = 0.030 and P = 0.046, respectively). On multivariate analysis, a significant difference was observed in patients with stage II cancer that was not observed in other stages.

Conclusions:  Nuclear PTEN expression gradually decrease during the normal–adenoma–adenocarcinoma–metastasis sequence, which suggests an important role for PTEN in carcinogenesis. Moreover, loss of nuclear PTEN expression was a marker of poor clinical outcome in patients with stage II colorectal cancer.
Histological assessment of pre‐transplant kidney biopsies is reproducible and representative
Authors: Snoeijs Maarten G J , Boonstra Liesbeth A , Buurman Wim A , Goldschmeding Roel , Van Suylen Robert Jan , Van Heurn L W Ernest , Peutz‐Kootstra Carine J
Published: December 2009

Snoeijs M G J, Boonstra L A, Buurman W A, Goldschmeding R, van Suylen R J, van Heurn L W E & Peutz‐Kootstra C J  (2010) Histopathology56, 198–202

Histological assessment of pre‐transplant kidney biopsies is reproducible and representative

Aims:  Histological examination of pre‐transplant renal biopsy specimens can be used to select grafts from older donors after cardiac death (DCD) with a satisfactory transplant outcome. The aim was to determine whether such biopsy specimens can be reproducibly scored between pathologists and are representative of the whole kidney.

Methods and results:  In renal biopsy specimens from DCD aged ≥60 years (n = 44), globally sclerosed glomeruli, vascular narrowing, tubular atrophy and interstitial fibrosis were scored by three independent pathologists according to the Pirani scoring system. Interobserver agreement on the sum of scores improved considerably with the introduction of a combined tubulo‐interstitial scoring system (intraclass correlation coefficient increased from 0.38 to 0.64). In small needle biopsy specimens (n = 144) obtained at autopsy, estimates of the proportion of globally sclerosed glomeruli were more precise with increasing sample size. Reasonably precise estimates may be obtained from specimens with at least seven glomeruli.

Conclusions:  It is feasible to implement pre‐transplant renal biopsy specimen analysis as a selection criterion in clinical practice in order to accept kidneys from marginal donors for transplantation.
Primary cutaneous leiomyosarcoma: clinicopathological analysis of 36 cases
Authors: Massi Daniela , Franchi Alessandro , Alos Llucia , Cook Martin , Di Palma Silvana , Enguita Ana B , Ferrara Gerardo , Kazakov Dmitry V , Mentzel Thomas , Michal Michal , Panelos John , Rodriguez‐Peralto José L , Santucci Marco , Tragni Gabrina , Zioga Aikaterini , Tos Angelo Paolo Dei
Published: December 2009

Massi D, Franchi A, Alos L, Cook M, Di Palma S, Enguita A B, Ferrara G, Kazakov D V, Mentzel T, Michal M, Panelos J, Rodriguez‐Peralto J L, Santucci M, Tragni G, Zioga A & Tos A P D (2010) Histopathology56, 251–262

Primary cutaneous leiomyosarcoma: clinicopathological analysis of 36 cases

Aims:  Cutaneous leiomyosarcomas (LMS) are rare in comparison with their deep‐seated soft tissue and uterine counterparts, and have been poorly characterized. The aim was to verify whether the clinical behaviour of purely dermal LMS is different from that of LMS with minimal subcutis invasion.

Methods and results:  Twenty‐one purely dermal LMS and 15 dermal LMS with minimal subcutis extension were analysed. Tumours developed in 27 men and nine women (age range 29–91 years); most tumours showed a fasciculated (n = 23), pilar‐type (n = 12) and pleomorphic (n = 1) pattern. During the follow‐up period (range 2–192, mean 41 months) recurrences occurred in 1/16 (6.2%) of tumours confined to the dermis and in 2/11 (18.1%) tumours with minimal subcutis extension. The three recurrent tumours were high‐grade LMS, two of which exhibited myxoid areas. One patient with a pleomorphic dermal LMS with minimal extension into fat developed distant metastases 15 years after diagnosis.

Conclusions:  For LMS involving the skin, it is advisable to recognize and indicate in the histopathology report the depth of dermal and/or subcutaneous extension, since even minimal subcutaneous involvement may be associated with late local recurrences and/or distant metastases, and therefore appropriate and long‐term follow‐up is needed.
High‐resolution array CGH of metanephric adenomas: lack of DNA copy number changes
Authors: Szponar Adrianna , Yusenko Maria V , Kovacs Gyula
Published: December 2009

Szponar A, Yusenko M V & Kovacs G (2010) Histopathology56, 212–216

High‐resolution array CGH of metanephric adenomas: lack of DNA copy number changes

Aims:  Previous karyotyping and fluorescence in situ hybridization analysis of metanephric adenomas (MAs) has yielded controversial data. The aim of this study was to detect small genomic alterations, if any, specific to MAs by applying high‐resolution oligoarrays.

Methods and results:  DNA extracted from paraffin blocks of six metanephric adenomas was hybridized onto Agilent oligoarrays with ∼43 000 in situ synthesized 60‐mer oligonucleotide probes that span coding and non‐coding sequences with an average spatial resolution of ∼35 kb. None of the metanephric adenomas showed DNA copy number changes. To confirm our results, DNA extracted from the paraffin block of a chromophobe renal cell carcinoma (RCC) was simultaneously hybridized to one of the four arrays on the same slides as an internal control. The chromophobe RCC showed loss of several chromosomes but no alteration was seen in MAs. We have confirmed the negative results by dye‐swap and sex mismatch hybridization experiments.

Conclusions:  Our high‐resolution oligoarray analysis indicates that metanephric adenomas lack DNA copy number alterations. This finding may help to differentiate between metanephric adenomas from Wilms’ tumour and papillary renal cell adenoma with overlapping phenotype.