bluuebunny/crossref_metadata_2025 huggingface dataset:
Galanin type 2 receptors regulate neuronal survival, susceptibility to seizures and seizure‐induced neurogenesis in the dentate gyrus
Authors: Mazarati Andrey , Lu Xiaoying , Kilk Kalle , Langel Ülo , Wasterlain Claude , Bartfai Tamas
Published: June 2004

The neuropeptide galanin has been implicated in inhibiting seizures and protecting hippocampal neurons from excitotoxic injury. In the hippocampus galanin acts through two receptor subtypes, GalR1, expressed in CA1, and GalR2, abundant in dentate gyrus. We developed an approach to induce and to study selective semichronic knockdown of GalR2 in the rat hippocampus. A 50% reduction of GalR2 binding was achieved by continuous infusion of complementary peptide nucleic acid antisense oligonucleotide into the dentate gyrus. This resulted in an increase in the severity of self‐sustaining status epilepticus induced by electrical stimulation of the perforant path, induced mild neuronal injury in the dentate hilus, augmented seizure‐induced hilar injury and inhibited seizure‐induced neurogenesis in the subgranular zone of the dentate gyrus. Our data suggest that in the dentate gyrus, galanin, acting through GalR2, inhibits seizures, promotes viability of hilar interneurons and stimulates seizure‐induced neurogenesis. These results are important for understanding the role of galanin and galanin receptor subtypes in the hippocampus both under normal conditions and in excitotoxic injury.
Transfer of plastid DNA from tobacco to the soil bacterium Acinetobacter sp. by natural transformation
Authors: De Vries Johann , Herzfeld Thilo , Wackernagel Wilfried
Published: June 2004

Acquisition of new genetic information by horizontal gene transfer is a major mechanism of genetic adaptation and evolution in prokaryotes. Naturally transformable cells of Acinetobacter sp. were exposed to plant DNA from leaf and root tissue of transplastomic tobacco. With the aadA gene (resistance against spectinomycin and streptomycin) as anchor se‐quence, the transfer of segments of the tobacco plastid DNA to Acinetobacter by homology‐facilitated illegitimate recombination occurred at a frequency of 1.2 × 10−7 per cell, which was about 0.1% of the frequency of fully homologous transfers. Without anchor sequence, transfer was not detected (≤1.3 × 10−10). The integrated plastid DNA segments extended up to 2539 nucleotides and often encompassed tobacco genes (trnL, ycf5). Expression of trnL (leucyl‐tRNA) in a transformant was shown by reverse transcription polymerase chain reaction. About 44% of integration events occurred at a single hot‐spot and 38% at other multiply used sites. All illegitimate recombination sites were GC‐rich microhomologies of 3–6 bp often neighboured by further microhomologies. The sites were located in plant DNA at the ends of distinct larger high‐GC regions, which suggests a role for GC‐aided association of heterologous sequences in illegitimate DNA end joining. The results show that integration of plant DNA into a bacterial genome by natural transformation is possible and is probably stimulated by hot‐spots of illegitimate recombination.
Isolation and characterisation of a class of carbohydrate oxidases from higher plants, with a role in active defence
Authors: Custers Jerôme H.H.V. , Harrison Stuart J. , Sela‐Buurlage Marianne B. , Van Deventer Els , Lageweg Wessel , Howe Peter W. , Van Der Meijs Pieter J. , Ponstein Anne S. , Simons Bert H. , Melchers Leo S. , Stuiver Maarten H.
Published: June 2004

In a search for novel plant‐derived antimicrobial proteins, we screened extracts from salicylic acid (SA)‐treated lettuce and sunflower leaves. These extracts displayed very potent antimicrobial activity against a set of phytopathogens. Characterisation of these extracts revealed that in both extracts, proteins of approximately 60 kDa were responsible for the antimicrobial activity. Further characterisation of these proteins and cloning of the respective cDNAs revealed close homology to a range of (plant) oxidases. Dissection of the enzymatic activity of both proteins revealed them to be carbohydrate oxidases (Helianthus annuus carbohydrate oxidase (Ha‐CHOX) and Lactuca sativa carbohydrate oxidase (Ls‐CHOX)) with broad substrate specificity and with hydrogen peroxide (H2O2) as one of the reaction products. The sunflower transcript, in addition to being SA inducible, was also inducible by fungal pathogens but not by ethylene and jasmonate. To determine whether Ha‐CHOX plays a role in pathogen defence, it was transformed into tobacco and the effect of resistance to Pectobacterium carotovorum ssp. carotovorum was examined. Transgenic plants overexpressing Ha‐CHOX displayed enhanced resistance to infection by this pathogen, and the resistance level was proportional to enzyme expression.
Disruption of the novel plant protein NEF1 affects lipid accumulation in the plastids of the tapetum and exine formation of pollen, resulting in male sterility in Arabidopsis thaliana
Authors: Ariizumi Tohru , Hatakeyama Katsunori , Hinata Kokichi , Inatsugi Rie , Nishida Ikuo , Sato Shusei , Kato Tomohiko , Tabata Satoshi , Toriyama Kinya
Published: June 2004

A novel male‐sterile mutant of Arabidopsis thaliana was isolated by means of T‐DNA tagging. Pollen abortion of the mutant was evident after microspore release, and pollen grains were completely absent at anthesis. Transmission electron microscope analysis revealed that primexine was coarsely developed, and that although sporopollenin was produced, it was not deposited onto the microspore plasma membrane. The sporopollenin that failed to be deposited aggregated and accumulated within the locule and on the locule wall. Finally, as no exine formation was observed, the mutant was named nef1. The plastoglobuli within the plastids of the tapetum were reduced, and lipid accumulation was considerably decreased. The mutant had a significantly altered leaf chloroplast ultrastructure and showed various growth defects. Lipid analysis revealed that the total lipid content in nef1 was lower than that in the wild type, which indicated that Nef1 was involved in lipid metabolism. Cloning of the full‐length Nef1 indicated that the gene encodes a novel plant protein of 1123 amino acids with limited sequence similarities to membrane proteins or transporter‐like proteins, and the NEF1 is predicted to be a plastid integral membrane protein. Motif analysis revealed that NEF1 contains prokaryotic membrane lipoprotein lipid attachment sites that are involved in maintaining cell envelope integrity. It is predicted that the Nef1 encodes a membrane protein that maintains the envelope integrity in the plastids.
Endocytosis against high turgor: intact guard cells of Vicia faba constitutively endocytose fluorescently labelled plasma membrane and GFP‐tagged K+‐channel KAT1
Authors: Meckel Tobias , Hurst Annette C. , Thiel Gerhard , Homann Ulrike
Published: June 2004

The relevance of endocytosis in plants against high turgor pressure has frequently been questioned on the basis of energetic considerations. Here, we examine the dynamics of the plasma membrane (PM) in turgid guard cells of Vicia faba by monitoring with confocal microscopy the fate of fluorescent styryl dyes (FM1‐43, FM2‐10 and FM4‐64). As a second marker, we also observe the retrieval of a fluorescent chimaera of the K+‐inward rectifying channel from Arabidopsis thaliana and the green fluorescent protein (KAT1::GFP). Analysis of cytoplasmic structures, which became labelled by the different styryl dyes, revealed that only FM4‐64, the most hydrophobic dye, was a reliable marker of endocytosis, whereas the two other styryl dyes resulted also in an unspecific labelling of different cytoplasmic structures including mitochondria. Over some minutes of incubation in continuous presence of these dyes, endocytic vesicles in the cortical cytoplasm beneath the PM were fluorescently labelled. The identification is based on the observation that the size distribution of these structures is very similar to that of endocytic vesicles obtained from patch‐clamp capacitance recordings. Also, these structures are frequently co‐labelled with KAT1::GFP. Taken together, the data show that turgid guard cells undergo vigorous constitutive endocytosis and retrieve membrane including the K+‐channel KAT1 from the PM via endocytic vesicles.
The calcium‐dependent protein kinase HvCDPK1 mediates the gibberellic acid response of the barley aleurone through regulation of vacuolar function
Authors: McCubbin Andrew G. , Ritchie Sian M. , Swanson Sarah J. , Gilroy Simon
Published: June 2004

In the aleurone cells of the cereal grain, gibberellic acid (GA) induces the secretion of hydrolases that mobilize endosperm reserves to fuel early seedling growth. GA is known to trigger a range of cellular responses, including increases in cytoplasmic calcium, vacuolar reserve mobilization, gene transcription, and the synthesis and secretion of hydrolases. To further define elements of the Ca2+‐dependent GA response machinery, we have cloned a Ca2+‐dependent protein kinase (HvCDPK1) from these cells. Although expression of an inactivated (D140N) version of this kinase did not affect GA‐induced gene expression or changes in cytosolic Ca2+, it did inhibit secretion, cell vacuolation, and vacuolar acidification, all responses linked to the GA response. Additionally, recombinant wild‐type HvCDPK1 activated the V‐type H+‐ATPase present in isolated aleurone vacuoles. These results suggest HvCDPK1 may mediate Ca2+‐dependent events of the GA response, such as control of vacuolar function, that lie downstream of transcriptional regulation.
Virus‐induced gene silencing in Solanum species
Authors: Brigneti Gianinna , Martín‐Hernández Ana M. , Jin Hailing , Chen Judy , Baulcombe David C. , Baker Barbara , Jones Jonathan D. G.
Published: June 2004

Virus‐induced gene silencing (VIGS) has been used routinely in Nicotiana benthamiana to assess functions of candidate genes and as a way to discover new genes required for diverse pathways, especially disease resistance signalling. VIGS has recently been shown to work in Arabidopsis thaliana and in tomato. Here, we report that VIGS using the tobacco rattle virus (TRV) viral vector can be used in several Solanum species, although the choice of vector and experimental conditions vary depending on the species under study. We have successfully silenced the phytoene desaturase (PDS) gene in the diploid wild species Solanum bulbocastanum and S. okadae, in the cultivated tetraploid S. tuberosum and in the distant hexaploid relative S. nigrum (commonly known as deadly nightshade). To test whether the system could be utilised as a rapid way to assess gene function of candidate resistance (R) genes in potato and its wild relatives, we silenced R1 and Rx in S. tuberosum and RB in S. bulbocastanum. Silencing of R1, Rx and RB successfully attenuated R‐gene‐mediated disease resistance and resulted in susceptible phenotypes in detached leaf assays. Thus, the VIGS system is an effective method of rapidly assessing gene function in potato.
Disruption of AtMRP4, a guard cell plasma membrane ABCC‐type ABC transporter, leads to deregulation of stomatal opening and increased drought susceptibility
Authors: Klein Markus , Geisler Markus , Suh Su Jeoung , Kolukisaoglu H. Üner , Azevedo Louis , Plaza Sonia , Curtis Mark D. , Richter Andreas , Weder Barbara , Schulz Burkhard , Martinoia Enrico
Published: June 2004

ATP‐binding cassette (ABC) transporters are membrane proteins responsible for cellular detoxification processes in plants and animals. Recent evidence shows that this class of transporters may also be involved in many other cellular processes. Because of their homology with human multidrug resistance‐associated proteins (MRP), cystic fibrosis transmembrane conductance regulator (CFTR) and sulfonylurea receptor (SUR), some plant ABC transporters have been implicated in the regulation of ion channel activities. This paper describes an investigation of the AtMRP4 gene and its role in stomatal regulation. Reporter gene studies showed that AtMRP4 is highly expressed in stomata and that the protein is localized to the plasma membrane. Stomatal aperture in three independent atmrp4 mutant alleles was larger than in wild‐type plants, both in the light and in the dark, resulting in increased water loss but no change in the photosynthetic rate. In baker's yeast, AtMRP4 shows ATP‐dependent, vanadate‐sensitive transport of methotrexate (MTX), an antifolate and a substrate of mammalian MRPs. Treatment with MTX reduced stomatal opening in wild‐type plants, but had no effect in atmrp4 mutants. These results indicate the involvement of AtMRP4 in the complex regulation of stomatal aperture.
The plant CDF family member TgMTP1 from the Ni/Zn hyperaccumulator Thlaspi goesingense acts to enhance efflux of Zn at the plasma membrane when expressed in Saccharomyces cerevisiae
Authors: Kim Donggiun , Gustin Jeffery L. , Lahner Brett , Persans Michael W. , Baek Dongwon , Yun Dae‐Jin , Salt David E.
Published: June 2004

To avoid metal toxicity, organisms have evolved mechanisms including efflux of metal ions from cells and sequestration into internal cellular compartments. Members of the ubiquitous cation diffusion facilitator (CDF) family are known to play an important role in these processes. Overexpression of the plant CDF family member metal tolerance protein 1 (MTP1) from the Ni/Zn hyperaccumulator Thlaspi goesingense (TgMTP1), in the Saccharomyces cerevisiaeΔ zinc resistance conferring (zrc)1Δ cobalt transporter (cot)1 double mutant, suppressed the Zn sensitivity of this strain. T. goesingense was found to contain several allelic variants of TgMTP1, all of which confer similar resistance to Zn in Δzrc1Δcot1. Similarly, MTP1 from various hyperaccumulator and non‐accumulator species also confer similar resistance to Zn. Δzrc1Δcot1 lacks the ability to accumulate Zn in the vacuole and has lower accumulation of Zn after either long‐ or short‐term Zn exposure. Expression of TgMTP1 in Δzrc1Δcot1 leads to further lowering of Zn accumulation and an increase in Zn efflux from the cells. Expression of TgMTP1 in a V‐type ATPase‐deficient S. cerevisiae strain also confers increased Zn resistance. In vivo and in vitro immunological staining of hemagglutinin (HA)‐tagged TgMTP1::HA reveals the protein to be localized in both the S. cerevisiae vacuolar and plasma membranes. Taken together, these data are consistent with MTP1 functioning to enhance plasma membrane Zn efflux, acting to confer Zn resistance independent of the vacuole in S. cerevisiae. Transient expression in Arabidopsis thaliana protoplasts also reveals that TgMTP1::green fluorescent protein (GFP) is localized at the plasma membrane, suggesting that TgMTP1 may also enhance Zn efflux in plants.
Floral induction in tissue culture: a system for the analysis of LEAFY‐dependent gene regulation
Authors: Wagner Doris , Wellmer Frank , Dilks Kieran , William Dilusha , Smith Michael R. , Kumar Prakash P. , Riechmann José Luis , Greenland Andrew J. , Meyerowitz Elliot M.
Published: June 2004

We have developed a versatile floral induction system that is based on ectopic overexpression of the transcription factor LEAFY (LFY) in callus. During shoot regeneration, flowers or floral organs are formed directly from root explants without prior formation of rosette leaves. Morphological and reporter gene analyses show that leaf‐like structures are converted to floral organs in response to LFY activity. Thus, increased levels of LFY activity are sufficient to bypass normal vegetative development and to direct formation of flowers in tissue culture. We found that about half of the cultured cells respond to inducible LFY activity with a rapid upregulation of the known direct target gene of LFY, APETALA1 (AP1). This dramatic increase in the number of LFY‐responsive cells compared to whole plants suggested that the tissue culture system could greatly facilitate the analysis of LFY‐dependent gene regulation by genomic approaches. To test this, we monitored the gene expression changes that occur in tissue culture after activation of LFY using a flower‐specific cDNA microarray. Induction of known LFY target genes was readily detected in these experiments. In addition, several other genes were identified that had not been implicated in signaling downstream of LFY before. Thus, the floral induction system is suitable for the detection of low abundance transcripts whose expression is controlled in an LFY‐dependent manner.
Nitric oxide attenuates ischaemia‐reperfusion (I/R) injury in the diabetic liver
Authors: Rogers Hollis , Zibari Gazi B , Roberts Jenifer , Turnage Richard , Lefer David J
Published: June 2004

Abstract:  Background:  Liver ischaemia‐reperfusion (I/R) occurs during resuscitation from haemorrhagic shock, hepatic transplantation and anatomic resection of the liver. This injury is associated with hepatocellular enzyme release and hepatocyte necrosis. The impact of chronic illnesses such as diabetes mellitus (DM) on hepatic I/R is unknown. This study determines the effect of DM on liver I/R using a murine model of type II DM in which the leptin receptor is defective. Preliminary studies suggest that animal models of DM have impaired endothelial nitric oxide (NO) release. Other studies suggest that NO attenuates hepatic I/R in phenotypically normal animals. We postulated that DM exacerbates hepatic I/R and that exogenous NO administration will attenuate hepatocellular injury.

Methods:  Non‐diabetic and diabetic (db/db) mice were anaesthetized and underwent laparotomy with the placement of a microvascular clip on the hepatic artery and portal vein supplying the medial and left lateral lobes of the liver rendering about 70% of the liver ischaemic. Hepatic ischaemia was maintained for 45 min after which time the clip was removed and the liver segments reperfused. The abdomen was closed and the animals maintained for 5 h of reperfusion. Hepatic injury was then assessed by measuring serum alanine and aspartate transaminases (ALT, AST) spectrophotometrically. Sections of liver reperfused for 24 h were stained with haematoxylin and eosin and the percentage of hepatocyte necrosis evaluated using morphometric techniques. Other animals undergoing hepatic I/R received the NO donor (DETA 100 µg/kg, i.v. 5 min prior to reperfusion). Time‐matched, sham‐operated animals served as controls. The data are expressed as mean ± SEM and analysed by ANOVA.

Results:  Serum AST and ALT levels were significantly higher in db/db animals vs. non‐diabetics, even in the absence of hepatic I/R (P < 0.01). Serum AST and ALT levels in db/db mice undergoing hepatic I/R were nearly five times greater than that of non‐diabetic animals (P < 0.01). Histologic examination of the livers of the diabetic animals undergoing I/R demonstrated significantly greater hepatocellular necrosis (zone III; 30–40%) when compared with non‐diabetic animals sustaining the same injury (zone III; 3–10%). The NO donor DETA totally prevented the increase in serum ALT and AST release associated with I/R in both the diabetic and non‐diabetic mice when compared with animals not receiving this agent (P < 0.01).

Conclusion:  This is the first study suggesting that DM exacerbates hepatic I/R and that NO donors will prevent this hepatocellular injury in the diabetic. Sixteen million Americans have DM. Understanding the effect of this chronic illness on the inflammatory response to injury is essential to improving clinical outcomes in these medically compromised patients.
Interstitial fluid analysis for assessment of organ function
Authors: Baicu Simona C , Simmons Patricia M , Campbell Lia H , Taylor Michael J , Brockbank Kelvin GM
Published: June 2004

Abstract:  Evaluation methods are required for non‐heart‐beating donor (NHBD) kidneys to ensure the success of transplantation. In this study, the microdialysis technique was employed for the ex‐vivo assessment of hypothermically preserved NHBD kidney function. Microdialysis probes were placed in the renal cortex of 2 h warm ischaemic porcine kidneys to monitor interstitial pyruvate dynamics during hypothermic machine perfusion with perfusate containing 29.4 mM fructose‐1,6‐diphosphate (FDP). The presence of exogenous FDP in the perfusate induced no changes in the renal flow rate and vascular resistance, renal artery effluent biochemistry, or pyruvate concentration relative to untreated control kidneys. Significant increases in pyruvate production (P < 0.05), however, were observed after 12 h of perfusion in the interstitial fluid of FDP‐treated kidneys relative to control kidneys. After 24 h of perfusion, interstitial fluid concentrations of pyruvate were 149.1 ± 58.4 vs. 55.6 ± 17.9 µM (P < 0.05) in the FDP and control group, respectively. The microdialysis probe collected the interstitial fluid directly from the cellular sites of metabolic and synthetic activity, where perfusate dilution was minimal. Consequently, the biochemical changes induced by the organ metabolic activity were detected only at the interstitial level, in the microdialysates. Interstitial fluid pyruvate may be a good indicator of kidney function. The addition of FDP to the perfusion solution during ischaemic kidney preservation resulted in enhanced pyruvate production in the extracellular space, indirectly reflecting an increase in anaerobic ATP production. The pyruvate will be transformed during organ reperfusion into acetyl Co‐A enzyme allowing an immediate start of aerobic metabolism. This in turn can increase the amount of ATP available to the cells and may help prevent reperfusion injury upon transplantation.
Impact of 24 h continuous hypothermic perfusion on heart preservation by assessment of oxidative stress
Authors: Fitton Torin P , Wei Chiming , Lin Ruxian , Bethea Brian T , Barreiro Christopher J , Amado Luciano , Gage Fred , Hare Joshua , Baumgartner William A , Conte John V
Published: June 2004

Abstract:  Introduction:  Despite investigating numerous solutions, additives, and techniques over the last two decades, extending donor heart preservation beyond 4–6 h has not been achieved. Hypothermic heart preservation (HP) induces oxidative stress (OS) with reactive oxygen species (ROS) production, causing DNA cleavage and impairing repair. Quantification of cardiomyocyte concentrations of DNA damage by‐products (8‐oxoG) and mismatch repair enzymes (MYH, OGG‐1, MSH2) reflects the severity of OS. If increased repair enzyme production is insufficient to repair injury, cell death occurs and functional outcomes are impacted. We investigated continuous hypothermic perfusion (CHP), a new form of HP, and the mechanism of injury associated with hypothermic storage, by assessing functional outcome and OS after allotransplantation of canine hearts.

Methods:  Fourteen canine hearts were harvested using standard techniques after baseline echocardiograms and haemodynamic parameters were obtained. The hearts were implanted after 24 h CHP (n = 10) or 4 h static preservation (SP; n = 4). After weaning from cardiopulmonary bypass (CPB), recipients were kept alive for 6 h. Repeat echocardiograms and haemodynamic parameters were obtained. Quantification of MYH, OGG‐1, and MSH2 concentrations were performed on biopsies using immunohistochemistry and Western blot analysis.

Results:  Twelve out of 14 hearts (8/10 CHP; 4/4 SP) were successfully weaned on moderate inotropic support. Post‐implant echocardiogram, completed in 6/10 CHP and 2/4 SP hearts, demonstrated hyperdynamic function and normal wall motion. The expression and activity of DNA repair enzymes was identical between normal baseline and CHP hearts.

Conclusion:  CHP reduces OS associated with prolonged hypothermic preservation and may allow longer preservation periods without compromising function. CHP offers several potential advantages: (1) resuscitation of non‐beating heart donor organs, (2) time for HLA tissue typing, (3) facilitate interventions improving graft function, and (4) increased organ sharing.
Post transplant lymphocele: a single centre experience
Authors: Atray Naveen K , Moore Frank , Zaman Fahim , Caldito Gloria , Abreo Kenneth , Maley Warren , Zibari G B
Published: June 2004

Abstract:  The occurrence of post renal transplant lymphocele is variable and the best approach to treatment is not well defined. The purpose of this study was to find out the incidence of post transplant lymphocele at our centre, identify demographic or surgical factors that may have influenced lymphocele formation, and distinguish the best approach to treatment. The charts of 138 consecutive renal transplant recipients from 1996 to 2001 were retrospectively reviewed. The demographic characteristics, comorbid illnesses, occurrence of lymphocele and its treatment modality were recorded. A total of 36 (26%) patients developed lymphoceles. There was a significant relationship between an increased body mass index (BMI) and lymphocele occurrence (P > 0.01). The recurrence rate with drainage alone was 33%, which decreased to 25% with sclerotherapy. In comparison, both laparoscopic and open surgical marsupialization had a much lower but similar recurrence rate of 12%. The laparoscopic method had less morbidity, a shortened hospital stay, and less infection than open surgery.
ABO blood group influences a candidate's likelihood of receiving an HLA zero antigen mismatch kidney
Authors: Bryan Christopher F , Cherikh Wida S , Cheng Yulin , Aeder Mark I , Muruve Nicolas A , Nelson Paul W , Shield Charles F , Warady Bradley A , Winklhofer Franz T
Published: June 2004

Abstract:  National sharing of HLA zero‐mismatched kidneys has improved long‐term graft survival. The distribution of those HLA‐matched kidneys by ABO blood group, however, has not been examined.

Utilizing the UNOS/OPTN (United Network for Organ Sharing/Organ Procurement Transplantation Network) database, we analysed 112 971 kidney waiting list registrations added during 6/3/95–31/12/00, and 8162 HLA zero‐mismatched (0 mm) primary kidney transplants in the USA during 1/1/88–31/3/02. We also analyzed A isoagglutinin titer histories for 87 blood group B end stage renal disease (ESRD) patients for whom at least 1 yr of testing was done. Blood group A patients received 40.1% of the HLA‐0 mm kidneys while having a 26.5% representation on the national waiting list. Blood group B patients comprised 17.4% of the waiting list, but received only 10.4% of the HLA‐0 mm kidneys. Most (89.6%) blood group B patients awaiting kidney transplantation have low levels of A isoagglutinins, making them eligible to receive a blood group A2 kidney transplant. The national HLA‐0 mm kidney allocation sharing system's imbalance by ABO blood group could be partially resolved in the future by allocating HLA‐0 mm blood group A2 kidneys to B patients.
Augmentation of T‐cell apoptosis by immunosuppressive agents
Authors: Takahashi K , Reynolds M , Ogawa N , Longo D L , Burdick J
Published: June 2004

Abstract:  The regulatory benefit of apoptosis (activation‐induced cell death, AICD) in T cells can be influenced by immunosupressive agents. We examined this for mycophenolate mofetile (MMF, using it's active metabolite, mycophenolate (MPA)) compared with rapamycin (RAPA) and the calcineurin inhibitors (CI) cyclosporin (CYA) and FK506 (FK). Pure T cells from peripheral blood leucocytes (PBL) were stimulated by anti‐CD3 plus anti‐CD28. Cell division (sequential cohort reduction in carboxyflourescein diacetate succinimidyl ester, CFSE) was used to measure proliferation and determine status of different cell generations without or with added drug at 4 d. Apoptosis was measured by Annexin V staining of activated cells using flow cytometry. We confirmed in this stringent system the inhibition of AICD by CI and showed that RAPA is intermediate and MPA most effective in this potentiation of AICD.
Update on the immunology, diagnosis and management of systemic lupus erythematosus
Authors: Reeves G. E. M.
Published: June 2004

Abstract

Lupus occurs with a prevalence of 2−9 in 10 000 people, targeting female and indigenous populations in particular. Lupus and related systemic autoimmune syndromes (scleroderma, Sjögren's syndrome, rheumatoid arthritis and polymyositis) result from a similar set of genetically and environmentally modulated immune disturbances, and the diagnostic and management approach to these conditions is broadly comparable. Evanescent , vague symptoms, restrictive diagnostic criteria and low diagnostic suspicion might have resulted in the under‐diagnosis of these problems in the past, imposing considerable burdens on sufferers and the community. Serological screening should be employed cautiously and wisely, given the rapidly occurring changes in methodology, which have lowered their specificity. Close liaison with the immunology laboratories performing these tests is therefore advisable. Clinicians should emphasize the improving prognosis of lupus and related conditions as a result of earlier disease detection, improved educational support systems and refined medical therapies. (Intern Med J 2004: 34: 338−347)
Holter monitoring in the diagnosis of stroke mechanism
Authors: Shafqat S. , Kelly P. J. , Furie K. L.
Published: June 2004

Abstract

Background: Atrial fibrillation (AF), an important and treatable cause of ischaemic stroke, can occur as a sustained or a paroxysmal arrhythmia. Continuous cardiac rhythm monitoring (Holter monitoring) is often performed in stroke patients to identify paroxysmal AF, which is an indication for warfarin anti­coagulation in this patient population.

Aim: The aim of this study was to assess the clinical utility of Holter monitoring in detecting occult AF in patients with possible cardioembolic stroke.

Methods: The medical records of ischaemic stroke patients consecutively hospitalized at a single academic centre during a one‐year period were reviewed. Data regarding patient demographics, stroke characteristics, electrocardiography and echocardiography results and duration and findings of Holter monitoring were abstracted. The primary outcome was yield of newly diagnosed AF on Holter monitoring.

Results: Of 465 consecutive patients admitted with a diagnosis of new ischaemic stroke, 210 underwent Holter monitoring. The mean duration of monitoring was 22.8 ± 4.0 h. Previously undiscovered AF was ­identified in five cases (2.4%), all of which represented non‐rheumatic AF. In three cases, the Holter test was negative despite AF documented on an admission electro­cardiogram.

Conclusions: Holter monitoring can identify occult ­paroxysmal AF, assisting targeted secondary prevention in patients with new ischaemic stroke. However, the standard 24‐h duration of monitoring probably under­estimates the prevalence of paroxysmal AF in this population. Prospective studies are indicated to evaluate the value of longer monitoring periods in stroke popu­lations. (Intern Med J 2004; 34: 305−309)
Primary immunodeficiency diseases in adulthood
Authors: Riminton D. S. , Limaye S.
Published: June 2004

Abstract

Occasionally, patients present with clinical episodes of infectious disease that seem to fall beyond the scope of normal immunocompetence. Examples include infections that are unusually persistent, recurrent or resistant to treatment, or those involving unexpected dissemination of disease or atypical pathogens. Recent cellular and molecular advances in immunology help to inform the investigation and management of these cases and provide opportunities for family surveillance and disease prevention. More than 100 separate primary (inherited) immunodeficiency disease (PID) states have now been defined, each with a range of syndromic manifestations. Many of these are rare diseases of childhood, but some, either in atypical or attenuated forms, emerge for the first time in adulthood. We outline a diagnostic approach to five groups of PID presenting in adulthood and describe the recently launched web‐based PID Register of Australia and New Zealand − a new mechanism for improving knowledge and service delivery to Australasian patients and their clinical carers. (Intern Med J 2004; 34: 348−354)
Vaccination for asthma exacerbations
Authors: Bardin P. G.
Published: June 2004

Abstract

Most asthma exacerbations are caused by common cold virus infections, predominantly rhinovirus infections. Full protection against repeat infections with the same rhinovirus serotype is given by serum neutralizing antibody, but cross‐reactive antibody developed against other serotypes could yield partial protection and result in attenuated cold and airway symptoms. It is proposed that vaccine‐mediated induction of cross‐reactive antibody might not prevent rhinovirus infections but might reduce severe asthma symptoms and exacerbations. (Intern Med J 2004; 34: 358−360)
Ethics as process and practice
Authors: Frank A. W.
Published: June 2004

Abstract

The noun form of ethics suggests something substantive; this substance is located in both offices of institutionalized experts and a body of canonical solutions to recognizable troubles. The present article recommends thinking of ethics not as substance but as a process of lives and decisions affecting each other over time. No single decision stands alone and no single patient can be considered as a stand‐alone entity. Examples include how people struggle with choices they should not have to make, how decisions at one time are affected by earlier decisions that are not considered within the purview of ethics, and how to deal with conflicts of legitimate interests. When institutional ethics becomes a prescribed activity, the goal of being ethical is easily forgotten. (Intern Med J 2004; 34: 355−357)
Impact of early screening for reflux in siblings on the detection of renal damage
Authors: Houle Anne‐Marie , Cheikhelard Alaa , Barrieras Diego , Rivest Marie‐Christine , Gaudreault Valérie
Published: June 2004

OBJECTIVE

To assess the impact of screening siblings after detecting significant vesico‐ureteric reflux (VUR) and renal scarring, as such screening might identify patients with VUR before urinary tract infections develop, but might also detect clinically insignificant VUR.

PATIENTS AND METHODS

We used a previously reported screening protocol to assess the clinical characteristics of patients, including the incidence of renal scarring, and their siblings, and compared the results. In all, 123 children were screened and 44 (36%) had VUR on voiding cystography. The median (range) age at screening was 9 (1–90) months.

RESULTS

The grades of VUR detected were < III in 61% and ≥ III in 39%; VUR was bilateral in 48%. In all, 37 siblings with VUR were assessed by ultrasonography; 70% were normal, including 12 (32%) children with VUR of grade ≥ III. When used, renal scintigraphy was normal in 74% of siblings, vs 18% of index patients. However, when screened after 2 years old, siblings had twice the risk of already having renal damage on renal scintigraphy (P = 0.04).

CONCLUSION

Early screening (≤ 2 years) appears to be more protective for avoiding renal damage than screening older patients. Thus we propose early screening in asymptomatic siblings to detect VUR before it becomes clinically significant.
Use of the holmium:YAG laser for ureterolithotripsy in children
Authors: Dogan Hasan S. , Tekgul Serdar , Akdogan Bulent , Keskin Mehmet S. , Sahin Ahmet
Published: June 2004

OBJECTIVE

To review our experience with rigid ureteroscopy and holmium:YAG laser for treating ureteric calculi in children.

PATIENTS AND METHODS

The study included 35 children who were treated with rigid ureteroscopy for ureteric calculi between November 1997 and June 2003 (15 boys and 20 girls; mean age 6.2 years, range 1–14). The mean (range) stone size was 8 (4–15) mm and the duration of anaesthesia 46.6 (15–90) min. The stone was in the distal third of the ureter in 33 children and in the proximal third in two. We used a 7.5/8/10 F rigid ureteroscopes with routine dilatation of the ureteric orifice. For lower ureteric stones, lithotripsy was carried out with holmium:YAG laser in 29 cases, a pneumatic impactor in two and forceps extraction in two. Both stones in the proximal ureter were pushed back into the collecting system. All the ureters were stented using JJ stents in 31 and ureteric catheters in four cases. The mean postoperative follow‐up was 12 (2–30) months.

RESULTS

Excluding the two stones pushed back, the stone‐free rate after a one‐stage procedure was 82% (27/33). With repeated procedures in the six (ESWL in two) remaining cases the success rate was 97% (32/33). The ureter was perforated in two patients within the first five in the series. There was no pyelonephritis or gross haematuria after surgery.

CONCLUSION

Ureteroscopy and lithotripsy using the holmium:YAG laser is effective and safe for treating ureteric stones in children, in experienced hands. The results would be even better using smaller and flexible ureteroscopes.
The role of the rhabdosphincter in female rat voiding
Authors: Streng Tomi , Santti Risto , Andersson Karl‐Erik , Talo Antti
Published: June 2004

OBJECTIVES

To obtain information on the mechanisms of female rat micturition using a model in which pressure was measured in the bladder and distal part of the urethra corresponding to the location of the rhabdosphincter, providing information on the role of the sphincter in opening and closing the urethral lumen.

MATERIALS AND METHODS

A micturition reflex was induced in adult anaesthetized (chloral hydrate and urethane) female rats by filling the bladder with saline. Bladder pressure (BP), urethral pressure (UP), electromyography (EMG) of the middle part of the rhabdosphincter, and urinary flow rate in the distal urethra were simultaneously recorded.

RESULTS

There were four phases of the micturition contraction, the second characterized by intraluminal pressure high‐frequency oscillations (IPHFOs) of BP. When a non‐oscillatory micturition contraction started, the BP increased and exceeded UP for the rest of the micturition contraction. Even though the BP increased during this first phase, the urethral lumen stayed closed. Its opening was indicated by a simultaneous decrease in BP and increase of UP as the fluid flowed from the bladder to the urethra. When the rhabdosphincter closed, as indicated by an EMG‐burst of the muscle, the UP declined, bladder pressure increased and the flow ceased. Because of momentary contractions of the rhabdosphincter, the UP and urine flow rate had the same periodicity as the IPHFOs of BP.

CONCLUSIONS

The simultaneous recording of the BP, UP, EMG of the rhabdosphincter and urinary flow rate showed the sequence of events during micturition. The rhabdosphincter acts as an ‘on‐off’ switch, causing interruptions in the urinary flow rate.
Interleukin‐2 blocks the antitumour activity caused by depletion of CD25+ cells in a murine renal adenocarcinoma model
Authors: Takeuchi Takumi , Konno‐Takahashi Naoko , Kasuya Yutaka , Ogushi Tetsuo , Nishimatsu Hiroaki , Kitamura Tadaichi
Published: June 2004

OBJECTIVE

To test the effectiveness of antimouse CD25 monoclonal antibody (mAb) against murine renal adenocarcinoma (RENCA) cells, as immunoregulatory/suppressor cells are known to be involved in tumour development in vivo, but the functions of these cells are not yet clear, and eliminating naive CD25 (interleukin‐2 receptor α)‐positive T cells elicits potent immune responses to syngeneic tumours in vivo.

MATERIALS AND METHODS

Aliquots of 1 × 104 or 1 × 105 RENCA cells were implanted into the subcapsule of the left kidney of syngeneic male Balb/c mice. Mice were injected with 125 µg of antimouse CD25 mAb to deplete CD25+ cells before RENCA implantation. Then 104 units of recombinant human interleukin‐2 (rhIL‐2) were subcutaneously injected twice daily for 7 days. Fourteen or 25 days later the tumour size was determined by laparotomy, and cells sorted using two‐colour flow cytometry.

RESULTS

Depletion of naive CD25+ cells with anti‐CD25 mAb and rhIL‐2 administration effectively induced anti‐RENCA tumour activity in Balb/c hosts. However, co‐administration of anti‐CD25 mAb and rhIL‐2 abrogated this significant suppression of RENCA tumour growth. RENCA implantation reduced the proportion of CD4+ cells among splenocytes, whereas anti‐CD25 mAb treatment increased it. The proportion of CD25+CD8+ cells among splenocytes and that of CD25+ cells among CD8+ cells were markedly reduced by co‐administration of anti‐CD25 mAb and rhIL‐2 with RENCA implantation. Both CD4+ and CD8+ cells were stained around the remnant microscopic RENCA tumour after anti‐CD25 mAb treatment.

CONCLUSION

Either depletion of naive CD25+ cells or rhIL‐2 administration suppressed RENCA tumour growth in murine hosts. However, co‐administration of anti‐CD25 mAb and rhIL‐2 abrogated this significant suppression of RENCA tumour growth.
Vaginal construction using sigmoid colon in children and young adults
Authors: Rajimwale Ashok , Furness Peter D. , Brant William O. , Koyle Martin A.
Published: June 2004

Authors from Denver describe their experience in neovaginal construction using sigmoid colon in young adults. They outline potential difficulties, particularly with patients having an android pelvis, but feel that the isolated sigmoid neovagina construction is widely applicable and can be used at any age. However, they point out that the long‐term satisfaction for intercourse needs long‐term evaluation.

The impact of early screening for reflux in siblings of those with VUR is evaluated by authors from Montreal. They found that screening at an early stage (<2 years) was more protective against renal damage than screening at a later stage. They suggested that the optimal way to proceed was early screening in asymptomatic siblings to detect VUR before it becomes clinically significant.

OBJECTIVE

To evaluate the age at which patients who required vaginal replacement (an uncommon procedure in children) were diagnosed, and the cause of their anomaly, and to relate these variables to the surgical outcome.

PATIENTS AND METHODS

Patients who had vaginal replacement at the author's institution between 1990 and 2002 were reviewed retrospectively. Depending on the age at reconstructive surgery, patients were divided into pre‐ and postpubertal groups.

RESULTS

A neovagina was constructed in 23 patients during the study period; sigmoid colon was used in 20 but not in two patients with cloacal exstrophy and in one with Mayer‐Rokitansky‐Kuster–Hauser syndrome (MRKHS). These cases were excluded from the analysis of outcomes and complications. Group 1 comprised patients diagnosed and treated before puberty and group 2 those diagnosed and/or treated afterward. In group 1 the presenting diagnoses included androgen insensitivity syndrome (AIS) in six patients, MRKHS in two, cloacal exstrophy in two, vaginal tumour in one, Müllerian duct renal aplasia cervicothoracic somite dysplasia, vertebral abnormalities, anal atresia, cardiac anomalies, tracheo‐oesophageal fistula, and/or oesophageal atresia, renal abnormalities and limb defects syndromes in one each. In group 2 the presenting diagnoses included MRKHS in seven, AIS in two, and congenital adrenal hyperplasia in one. Complications included superficial wound infection (two patients), recurrent introital stenosis, and blind loop mucocele, complete stenosis of perineal neovaginal opening (one each) and dyspareunia in three. Neither age nor pelvic habitus (android vs gynaecoid) influenced the outcome, and the cosmetic results were excellent in all the patients.

CONCLUSION

Isolated sigmoid neovaginal construction appears to be applicable to many diagnoses and in patients at any age. Although an android pelvis can present technical challenges, in this experience it was not associated with a greater complication rate. The long‐term satisfaction with the sigmoid neovagina for intercourse, especially in those constructed before puberty, still requires long‐term evaluation.
Laparoscopic radical prostatectomy: current status
Authors: Hasan Waleed A. , Gill Inderbir S.
Published: June 2004

The role of laparoscopic radical prostatectomy is reviewed by authors from the Cleveland clinic. They discuss not only its history and development but also its current status in the management of prostate cancer. Bladder cancer is reviewed comprehensively by authors from Miami and Iowa; not only TCC but also the rather uncommon small cell neoplasm.

Finally, authors from London assess the use of tissue microarrays in the validation of markers which may have diagnostic, prognostic and therapeutic significance in urological oncology.
Consensus statement on bladder injuries
Authors: Gomez Reynaldo G. , Ceballos Lily , Coburn Michael , Corriere Joseph N. , Dixon Christopher M. , Lobel Bernard , McAninch Jack
Published: June 2004

The consensus on genitourinary trauma continues this month with the statement on bladder trauma from several internationally recognised experts on the subject. They describe blunt, penetrating and iatrogenic injuries and their management, considering paediatric injuries separately. They underline the importance of prompt diagnosis and treatment, stressing that problems raised when the diagnosis is delayed.
Matched‐cohort analysis of patients with prostate cancer followed with observation or treated with three‐dimensional conformal radiation therapy
Authors: Kramer Noel M. , Horwitz Eric M. , Uzzo Robert G. , Hanlon Alexandra L. , Hanks Gerald E.
Published: June 2004

OBJECTIVES

To compare the outcome of similar patients with prostate cancer treated by either observation or three‐dimensional conformal radiation therapy (3‐DCRT).

PATIENTS AND METHODS

The study included 69 patients with nonmetastatic prostate cancer who were observed only; the indications included indolent disease, significant medical comorbidities and refusal of treatment. Of these, 62 patients had palpable T1–T2a and seven T2b–T3a disease, a median Gleason score of 6 and a median initial prostate‐specific antigen (PSA) level of 5.3 ng/mL. A matched‐cohort analysis of 69 patients, based on palpation T category, Gleason score and initial PSA, was used to compare the outcome between the observation and 3‐DCRT groups. The median radiation dose for latter was 72 Gy.

RESULTS

The median follow‐up for the observed patients was 49 months. The 5‐ and 8‐year actuarial rates of freedom from distant metastases were 100% and 93%, respectively, and the actuarial overall survival rates 94% and 73%, respectively. Seven observed patients had local disease progression on physical examination. Four patients who initially were observed received radiation therapy later for a rising PSA and/or local disease progression. For the 69 matched 3‐DCRT patients, the overall 5‐year rate for no biochemically evident disease was 74%. The respective 5‐ and 8‐year actuarial rates of freedom from distant metastases were 95% and 95%, and actuarial overall survival rates 95% and 75%. There were no significant differences in distant metastasis and overall survival rates between the groups, and no deaths from prostate cancer in either group.

CONCLUSIONS

Observation is a reasonable alternative to treatment in selected patients. During the 5‐year follow‐up the progression rates were relatively low, and there was no difference in distant metastasis or overall survival between the groups. As the follow‐up was short a longer follow‐up is needed to determine whether the outcome of those patients who chose observation will remain comparable to that in those undergoing immediate 3‐DCRT.