bluuebunny/crossref_metadata_2025 huggingface dataset:
Human Werner helicase interacting protein 1 (WRNIP1) functions as a novel modulator for DNA polymerase δ
Authors: Tsurimoto Toshiki , Shinozaki Ayako , Yano Masaki , Seki Masayuki , Enomoto Takemi
Published: January 2005

Human WRNIP1, a Werner DNA helicase interacting protein 1, was expressed in insect cells and E. coli. The purified protein behaved as a homo‐oligomeric complex with a native molecular mass indicative of an octamer, and the complex copurified with an ATPase activity that was stimulated by double‐stranded DNA ends. As suggested by genetic studies of budding yeast WRNIP1/Mgs1, the purified human WRNIP1 complex interacted physically with human DNA polymerase δ (pol δ), stimulating its DNA synthesis activity more than fivefold in the presence or absence of proliferating cell nuclear antigen. Analysis of reaction products demonstrated the stimulation to be partly due to an increased processivity of pol δ but more importantly to an increase in its initiation frequency. Addition of ATP to reactions partially suppressed stimulation by WRNIP1. Furthermore, a mutant WRNIP1 lacking ATPase activity could stimulate pol δ normally but was insensitive to suppression by ATP. These results indicate that WRNIP1 functions as a modulator for initiation or restart events during pol δ‐mediated DNA synthesis and that its ATPase activity is utilized to sense DNA ends and to regulate the extent of stimulation.
The Drosophila Toucan protein is a new mitotic microtubule‐associated protein required for spindle microtubule stability
Authors: Mirouse Vincent , Dastugue Bernard , Couderc Jean‐Louis
Published: January 2005

Mitotic spindle dynamics are highly dependent on proteins that interact with microtubules to influence their organization or stability. Here, we show that the Drosophila Toucan protein interacts directly with microtubules. Its localization to the microtubule network when it is expressed in mammalian cells and its direct interaction with microtubules in vitro are dependent on its central basic domain. Moreover, Toc expression in mammalian cells strongly protects microtubules from depolymerization. By using in vivo inducible RNAi in syncytial embryos, we generated a dose‐sensitive loss of function of toucan, demonstrating that this technique is an efficient method for inactivating a maternal transcript. This enabled us to accurately characterize several new mitotic defects from the early to the late phases of mitosis, depending on Toucan depletion level. Toucan is required for metaphase spindle formation and centrosome anchoring to the poles. Then, during anaphase, Toc depletion affects kinetochore microtubules and therefore chromosome segregation. Toc is also necessary for central spindle formation by the interpolar microtubules. In contrast, astral microtubules are not disturbed by Toc depletion. Taken together, our results show that Toucan is a microtubule‐associated protein specifically required for the stability of spindle microtubules throughout mitosis.
Transgenic over‐expression of GATA‐1 mutant lacking N‐finger domain causes hemolytic syndrome in mouse erythroid cells
Authors: Nakano Mayu , Ohneda Kinuko , Yamamoto‐Mukai Harumi , Shimizu Ritsuko , Ohneda Osamu , Ohmura Sakie , Suzuki Mikiko , Tsukamoto Saho , Yanagawa Toru , Yoshida Hiroshi , Takakuwa Yuichi , Yamamoto Masayuki
Published: January 2005

Transcription factor GATA‐1 is essential for erythroid cell differentiation. GATA‐binding motifs have been found in the regulatory regions of various erythroid‐specific genes, suggesting that GATA‐1 contributes to gene regulation during the entire process of erythropoiesis. A GATA‐1 germ‐line mutation results in embryonic lethality due to defective primitive erythropoiesis and GATA‐1‐null embryonic stem cells fails to differentiate beyond the proerythroblast stage. Therefore, the precise roles of GATA‐1 in the later stages of erythropoiesis could not be clarified. Under the control of a GATA‐1 gene hematopoietic regulatory domain, a GATA‐1 mutant lacking the N‐finger domain (ΔNF mutant) was over‐expressed in mice. These mice exhibited abnormal morphology in peripheral red blood cells (RBCs), reticulocytosis, splenomegaly, and erythroid hyperplasia, indicating compensated hemolysis. These mice were extremely sensitive to phenylhydrazine (PHZ), an agent that induces hemolysis, and their RBCs were osmotically fragile. Importantly, the hemolytic response to PHZ was partially restored by the simultaneous expression of wild‐type GATA‐1 with the ΔNF mutant, supporting our contention that ΔNF protein competitively inhibits the function of endogenous GATA‐1. These data provide the first in vivo evidence that the NF domain contributes to the gene regulation that is critical for differentiation and survival of mature RBCs in postnatal erythropoiesis.
Crystal structure of the protein histidine phosphatase SixA in the multistep His‐Asp phosphorelay
Authors: Hamada Keisuke , Kato Masato , Shimizu Toshiyuki , Ihara Kentaro , Mizuno Takeshi , Hakoshima Toshio
Published: January 2005

The multiple histidine‐aspartate phosphorelay system plays a crucial role in cellular adaptation to environments in microorganisms and plants. Like kinase‐phosphatase systems in higher eukaryotes, the multiple steps provide additional regulatory checkpoints with phosphatases. The Escherichia coli phosphatase SixA exhibits protein phosphatase activity against the histidine‐containing phosphotransfer (HPt) domain located in the C‐terminus of the histidine kinase ArcB engaged in anaerobic responses. We have determined the crystal structures of the free and tungstate‐bound forms of SixA at 2.06 Å and 1.90 Å resolution, respectively. The results provide the first three‐dimensional view of a bacterial protein histidine phosphatase, revealing a compact α/β architecture related to a family of phosphatases containing the arginine‐histidine‐glycine (RHG) motif at their active sites. Compared with these RHG phosphatases, SixA lacks an extra α‐helical subdomain as a lid over the active site, thereby forming a relatively shallow groove important for the accommodation of the HPt domain of ArcB. The tungstate ion, which mimics the substrate phosphate group, is located at the centre of the active site where the active residue, His8, points to the tungsten atom in the mode of in‐line nucleophilic attack.
Haptoglobin dampens endotoxin‐induced inflammatory effects both in vitro and in vivo
Authors: Arredouani Mohamed S. , Kasran Ahmad , Vanoirbeek Jeroen A. , Berger Frank G. , Baumann Heinz , Ceuppens Jan L.
Published: January 2005

We report that haptoglobin, an acute‐phase protein produced by liver cells in response to interleukin‐6 (IL‐6), can modulate the inflammatory response induced by endotoxins. We provide evidence that haptoglobin has the ability to selectively antagonize lipopolysaccharide (LPS) effects in vitro by suppressing monocyte production of tumour necrosis factor‐α, IL‐10 and IL‐12, while it fails to inhibit the production of IL‐6, IL‐8 and IL‐1 receptor antagonist. In two animal models of LPS‐induced bronchopulmonary hyperreactivity and endotoxic shock, haptoglobin knockout mice were more sensitive to LPS effects compared to their wild‐type counterparts. The present data suggest that haptoglobin regulates monocyte activation following LPS stimulation. The increase in haptoglobin levels during an acute‐phase reaction may generate a feedback effect which dampens the severity of cytokine release and protects against endotoxin‐induced effects.
Comparison of the expressed porcine Vβ and Jβ repertoire of thymocytes and peripheral T cells
Authors: Butler John E. , Wertz Nancy , Sun Jishan , Sacco Randy E.
Published: January 2005

Transcripts of more than 300 unique T‐cell receptor‐β (TCR‐β) V‐D‐J rearrangements recovered from porcine thymocytes and peripheral T cells were compared. We identified 19 groups (families) of porcine Vβ genes in seven supergroups and provisionally named 17 groups based on their sequence similarity with recognized human Vβ gene families. TRBV4S, 5S, 7S and 12S accounted for > 80% of all Vβ usage, and usage of these groups by thymocytes and peripheral T cells was highly correlated. No TRBV group was uniquely expressed in significant numbers in thymocytes, although small numbers of TRBV groups 2S, 9S and 15S were only recovered from T cells. Usage of Jβ segments from the 5′ D‐J‐C duplicon in thymocytes and peripheral T cells directly correlated with their 5′ position in the locus, and Jβ1·1, 1·2 and 1·3 accounted for ≥ 35% of all Jβ usage in both cell types. This contrasts with the usage of Jβ2 segments in that Jβ2·4, 2·5 and 2·7 accounted for ≈ 30% of Jβ usage by T cells and thymocytes. Jβ2·7 was threefold more frequent among T cells than thymocytes. The Vβ/Jβ combination was not random. Jβ1·1 and 1·2 were used in 29% of rearrangements with high frequency among the major Vβ groups. Combinations of TRBV4 and V12 with Jβ2·7 were only found in T cells and accounted for half of all Jβ2·7 usage. These studies show that unlike porcine heavy chain VH genes, the occurrence and relative usage of porcine TCR‐Vβ groups resembles that of humans. Thus, highly related gene systems can individually diverge within a species.
Follicular dendritic cell dedifferentiation reduces scrapie susceptibility following inoculation via the skin
Authors: Mohan Joanne , Bruce Moira E. , Mabbott Neil A.
Published: January 2005

Transmissible spongiform encephalopathies (TSEs) are a group of subacute infectious neurodegenerative diseases that are characterized by the accumulation in affected tissues of PrPSc, an abnormal isoform of the host prion protein (PrPc). Following peripheral exposure, TSE infectivity and PrPSc usually accumulate in lymphoid tissues prior to neuroinvasion. Studies in mice have shown that exposure through scarified skin is an effective means of TSE transmission. Following inoculation via the skin, a functional immune system is critical for the transmission of TSEs to the brain, but until now, it has not been known which components of the immune system are required for efficient neuroinvasion. Temporary dedifferentiation of follicular dendritic cells (FDCs) by treatment with an inhibitor of the lymphotoxin‐β receptor signalling pathway (LTβR‐Ig) 3 days before or 14 days after inoculation via the skin, blocked the early accumulation of PrPSc and TSE infectivity within the draining lymph node. Furthermore, in the temporary absence of FDCs before inoculation, disease susceptibility was reduced and survival time significantly extended. Treatment with LTβR‐Ig 14 days after TSE inoculation also significantly extended the disease incubation period. However, treatment 42 days after inoculation did not affect disease susceptibility or survival time, suggesting that the infection may have already have spread to the nervous system. Together these data show that FDCs are essential for the accumulation of PrPSc and infectivity within lymphoid tissues and subsequent neuroinvasion following TSE exposure via the skin.
Human monocyte isolation methods influence cytokine production from in vitro generated dendritic cells
Authors: Elkord Eyad , Williams Paul E. , Kynaston Howard , Rowbottom Anthony W.
Published: January 2005

There is growing interest in the in vitro generation of dendritic cells (DC) from peripheral blood monocytes, but the effect of the method chosen to isolate CD14+ monocytes for subsequent DC generation is poorly documented. The method used to isolate monocytes may have an impact on the subsequent function of DC by affecting their ability to express costimulatory molecules (CD80/86), maturation marker (CD83) and/or to produce important immunomodulatory cytokines. In this study, we show that the positive selection of monocytes by anti‐CD14‐coated microbeads inhibits the lipopolysaccharide (LPS)‐induced production of interleukin (IL)‐12, IL‐10 and tumour necrosis factor‐α (TNF‐α) from human DC. However, when DC were grown from monocytes isolated by plastic adherence, LPS induced the production of much higher levels of these cytokines. DC derived from adherence‐isolated monocytes induced the development of potent cytotoxic T lymphocytes of the Tc1 subset specific for influenza matrix protein, as confirmed by interferon‐γ (IFN‐γ) enzyme‐linked immunosorbent spot‐forming cell assay (ELISPOT), cytotoxicity assay, major histocompatibility complex (MHC)–peptide tetrameric complexes and T helper 1/T helper 2 (Th1/Th2) cytokine production assays.
Tick saliva inhibits differentiation, maturation and function of murine bone‐marrow‐derived dendritic cells
Authors: Cavassani Karen A. , Aliberti Júlio C. , Dias Alexandra R. V. , Silva João S. , Ferreira Beatriz R.
Published: January 2005

Haematophagous arthropod vectors such as mosquitoes, tsetse flies, sandflies and ticks have evolved salivary immunomodulatory factors that prevent the vertebrate host from rejecting them meanwhile enhancing pathogen transmission. As dendritic cells (DC) play a major role in host immune responses, we studied the effects of Rhipicephalus sanguineus tick saliva on DC differentiation and maturation. Flow cytometry analysis revealed that the addition of saliva to bone marrow cells inhibits the differentiation of DC and decreased the population of differentiated immature DC, increasing the levels of major histocompatibility complex (MHC) class II while not altering the expression of costimulatory (CD40, CD80 and CD86) and adhesion (CD54) molecules. Furthermore, maturation of DC stimulated by lipopolysaccharide (LPS) in the presence of saliva resulted in a lower expression of costimulatory molecules, but did not alter the up‐regulation of MHC class II and CD54. The lipopolysaccharide (LPS)‐matured DC cultured with saliva also presented reduced production of interleukin‐12, whereas interleukin‐10 production was unaltered. Assessment of the function of DC cultured with tick saliva revealed them to be poor stimulators of cytokine production by antigen‐specific T cells. Our data indicate a novel modulatory role for the saliva of arthropod vectors at an initial step of the immune response through the inhibition of differentiation and maturation of DC into functional antigen‐presenting cells.
Mucosal delivery of CpG oligodeoxynucleotides expands functional dendritic cells and macrophages in the vagina
Authors: Sajic Dusan , Patrick Amy J. , Rosenthal Kenneth L.
Published: January 2005

Antigen‐presenting cells (APC) are specialized sentinel cells that sense pathogens within tissues and then activate appropriate immune effector cells in lymphoid organs. Recent evidence, however, suggests that APC can also induce effector cells in non‐lymphoid organs. The purpose of this study was to determine the effect of intravaginal (IVAG) delivery of CpG‐oligodeoxynucleotide (ODN) on expansion of resident genital APC. Our results show that delivery of CpG‐ODN to the murine genital tract induced a rapid and significant, but transient expansion of genital APC in situ. As early as 12 hr post CpG‐ODN delivery, we observed an enhanced level of F4/80+ major histocompatibility complex (MHC) class II‐negative macrophages in the genital tissue. This was followed by increased levels of F4/80/MHC class II double‐positive cells, as well as MHC class II, CD11c and CD86 triple‐positive dendritic cells (DC) at 48 hr. Expanded APC levels at 48 hr post CpG‐ODN resulted in increased ability of genital cells to induce an allogenic mixed leucocyte reaction. By 72 hr after CpG‐ODN treatment, APC levels were not distinguishable from naïve levels. Therefore, these results clearly show that administration of CpG‐ODN to the genital tract induced a marked but transient enhancement of APC within the genital tissue, and that these APC appear to possess functional capacity. Furthermore, these results indicate that IVAG–CpG‐ODN may be an important factor for the enhancement of local antigen presentation in the genital tract through increased DC numbers.
Major histocompatibility class II molecules prevent destructive processing of exogenous peptides at the cell surface of macrophages for presentation to CD4 T cells
Authors: Von Delwig Alexei , Musson Julie A. , Gray Joe , Mckie Norman , Robinson John H.
Published: January 2005

We studied factors affecting major histocompatibility complex class II (MHC‐II)‐restricted presentation of exogenous peptides at the surface of macrophages. We have previously shown that peptide presentation is modulated by surface‐associated proteolytic enzymes, and in this report the role of the binding of MHC‐II molecules in preventing proteolysis of exogenous synthetic peptides was addressed. Two peptides containing CD4 T‐cell epitopes were incubated with fixed macrophages expressing binding and non‐binding MHC‐II, and supernatants were analysed by high‐performance liquid chromatography and mass spectrometry to monitor peptide degradation. The proportion of full‐length peptides that were degraded and the number of peptide fragments increased when non‐binding macrophages were used, leading to reduction in peptide presentation. When MHC‐II molecules expressed on the surface of fixed macrophages were blocked with monoclonal antibody and incubated with peptides and the supernatants were transferred to fixed macrophages, a significant reduction in peptide presentation was observed. Peptide presentation was up‐regulated at pH 5·5 compared to neutral pH, and the latter was found to be the pH optimum of the proteolytic activity of the surface enzymes involved in the degradation of exogenous peptides and proteins. The data suggest that MHC‐II alleles that bind peptides protect them from degradation at the antigen‐presenting cell surface for presentation to CD4 T cells and we argue that this mechanism could be particularly pronounced at sites of inflammation.
Glycation of CD59 impairs complement regulation on erythrocytes from diabetic subjects
Authors: Davies Catherine S. , Harris Claire L. , Morgan B. Paul
Published: January 2005

Type 1 diabetes is associated with anaemia. Although the underlying mechanisms remain unclear, the accompanying reticulocytosis implies that erythrocyte lifespan in the circulation is shortened. Among the factors that permit prolonged survival of erythrocytes are the membrane complement regulators. In conditions such as paroxysmal nocturnal haemoglobinuria, where erythrocyte expression of these regulators is reduced, erythrocyte survival is compromised and anaemia follows. Recent in vitro evidence indicates that one of the key membrane complement regulators, CD59, is inactivated by glycation in the presence of high concentrations of glucose or other glycating sugars. To ascertain whether glycation‐induced inactivation of CD59 occurrs in vivo we examined CD59 surface expression and function on erythrocytes from a cohort with poorly controlled type 1 diabetes (hyperglycaemic) and from matched normoglycaemic controls. Although expression of CD59, assessed using polyclonal anti‐CD59 antiserum, was similar in the two groups, erythrocytes from hyperglycaemic individuals were more susceptible to lysis by complement, entirely as a result of the loss of functional CD59. These data implicate glycation‐induced inactivation of CD59 as a factor contributing to anaemia in type 1 diabetes.
Interleukin‐24 and its receptors
Authors: Wang Mai , Liang Peng
Published: January 2005

Interleukin 24 (IL‐24) is a new member of the IL‐10 family of cytokines and it signals through two heterodimeric receptors: IL‐20R1/IL‐20R2 and IL‐22R1/IL‐20R2. Upon binding to its receptors, IL‐24 induces rapid activation of Stat‐1 and Stat‐3 transcription factors, which appear to play a role in cell survival and proliferation. Under physiological conditions, the major sources of IL‐24 are the activated monocytes and T helper 2 cells, whereas the major IL‐24 target tissues, based on the receptor expression pattern, are non‐haematopoietic in origin, and include skin, lung and reproductive tissues. Structurally and functionally, IL‐24 is highly conserved across species. This review highlights our current knowledge of IL‐24 as a cytokine, with much less emphasis placed on the non‐receptor‐mediated functions (a subject of several reviews) focused on in much of the earlier literature on IL‐24. The potential roles of IL‐24 as part of a complex cytokine network in wound healing, psoriasis and cancer are discussed.
Expression and possible functions of DNA lesion bypass proteins in spermatogenesis
Authors: VAN DER LAAN ROALD , BAARENDS WILLY M. , WASSENAAR EVELYNE , ROEST HENK P. , HOEIJMAKERS JAN H. J. , GROOTEGOED J. ANTON
Published: January 2005

In mammalian cells, there is a complex interplay of different DNA damage response and repair mechanisms. Several observations suggest that, in particular in gametogenesis, proteins involved in DNA repair play an intricate role in and outside the context of DNA repair. Here, we discuss the possible roles of proteins that take part in replicative damage bypass (RDB) mechanisms, also known as post‐replication DNA repair (PRR), in germ line development. In yeast, and probably also in mammalian somatic cells, RDB [two subpathways: damage avoidance and translesion synthesis (TLS)] prevents cessation of replication forks during the S phase of the cell cycle, in situations when the replication machinery encounters a lesion present in the template DNA. Many genes encoding proteins involved in RDB show an increased expression in testis, in particular in meiotic and post‐meiotic spermatogenic cells. Several RDB proteins take part in protein ubiquitination, and we address relevant aspects of the ubiquitin system in spermatogenesis. RDB proteins might be required for damage avoidance and TLS of spontaneous DNA damage during gametogenesis. In addition, we consider the possible functional relation between TLS and the induction of mutations in spermatogenesis. TLS requires the activity of highly specialized polymerases, and is an error‐prone process that may induce mutations. In evolutionary terms, controlled generation of a limited number of mutations in gametogenesis might provide a mechanism for evolvability.
Seminal Factor VIII and von Willebrand Factor: a possible role of the conventional clotting system in human semen?
Authors: LWALEED BASHIR A. , GREENFIELD ROBERT , ROYLE ERIC , Birch Brian , Cooper Alan J.
Published: January 2005

Factor (F) VIII circulates in blood complexed with von Willebrand Factor (vWF). Deficiency or defect accounts for haemophilia A and vWF disease. In blood, FVIII functions as a co‐factor for FIXa in the activation of FX. Human semen coagulates and liquefies in a process that resembles and has some links with the conventional haemostatic process. A study elsewhere has detected traces, but not measurable levels, of FVIII coagulant activity (FVIII:C). In the present study we have assessed FVIII antigen (FVIII:Ag), FVIII:C and vWF antigen (vWF:Ag) levels in 159 semen specimens obtained from sub‐fertile (n = 21), normally fertile (n = 38), fertile donors (n = 32), and vasectomized men (n = 57). Seminal FVIII:Ag levels were also measured in a group defined by several parameters derived from the World Health Organization (WHO) fertility criteria, termed ‘pooled normal semen parameters’ (PNSP). Factor VIII:Ag levels were compared with conventional fertility parameters. In addition, both FVIII:C and vWF:Ag were assessed in a separate group of normal individuals (n = 11). Factor VIII:Ag, FVIII:C and vWF were present and quantifiable in human semen. Factor VIII:Ag levels were significantly lower in vasectomy subjects compared with donors (p = 0.01) or PNSP group (p = 0.01). Several trends taken together suggest an associations between FVIII:Ag and semen quality. Parallel investigations demonstrate FV, FVII, FVIIa, FIX, FIXa, FXa, FXI, FXII, tissue factor (TF) and tissue factor pathway inhibitor (TFPI) in semen. The present report therefore provides further evidence for the presence of a functioning clotting system in human semen.
Intracerebroventricular Injections of Prolactin Counteract the Antagonistic Effect of Bromocriptine on Rabbit Maternal Behaviour
Authors: González‐Mariscal G. , Chirino R. , Flores‐Alonso J. C. , Rosenblatt J. S. , Beyer Carlos
Published: January 2005

To investigate the participation of prolactin in nest‐building and maternal behaviour in rabbits, we administered (from pregnancy day 26 to parturition) rabbit prolactin (rbPRL; or vehicle) intracerebroventricularly (i.c.v.) to primiparous animals injected with bromocriptine subcutaneously (s.c.). Control females (given vehicle s.c. and i.c.v.) built a maternal nest (of straw and body hair) in 77% of cases. This proportion decreased to 19% in the bromocriptine‐only group (P < 0.05) and increased to 93% in the group given bromocriptine plus rbPRL (P > 0.05). Maternal behaviour (i.e. the adoption of a crouching posture over the litter inside the nest box) was expressed by 77% of control rabbits, 19% of bromocriptine‐only animals (P < 0.05) and 57% of females given bromocriptine plus rbPRL (P > 0.05). Values of nonmaternal activities (i.e. scent‐marking, ambulation in an open field) were similar among the three studied groups. These results suggest that prolactin, acting in late pregnancy, plays a major role in the stimulation of nest‐building and maternal behaviour in rabbits.
Plasma and Hypothalamic Peptide‐Hormone Levels Regulating Somatotroph Function and Energy Balance in Fed and Fasted States: A Comparative Study in Four Strains of Rats
Authors: Kappeler L. , Zizzari P. , Grouselle D. , Epelbaum J. , Bluet‐Pajot M. T.
Published: January 2005

Both growth hormone (GH)/insulin growth factor (IGF)‐1 axis and energy balance have been implicated in longevity independently. The aim of the present study was to characterize the effect of a 72‐h fasting period at 3 months of age in four different rat strains: (i) Wistar and (ii) Fischer 344 rats, which develop obesity with age, and (iii) Brown Norway and (iv) Lou C rats, which do not. Wistar rats ate more, were significantly bigger, and presented with higher plasma leptin and lower ghrelin levels and hypothalamic growth hormone‐releasing hormone (GHRH) content than rats from the three other strains. Plasma insulin and IGF‐1 levels were lower in Brown Norway and Lou C rats, and somatostatin content was lower in Brown Norway rats only. Glycaemia was lower in Lou C rats that displayed a lower relative food intake compared to Fischer and Wistar rats. Brown Norway rats showed a greater caloric efficiency than the three other strains. Concerning major hypothalamic neuropeptides implicated in feeding, similar amounts were detected in the four strains for neuropeptide Y, agouti‐related peptide, galanin, melanin‐concentrating hormone, α‐melanocortin‐stimulating hormone (α‐MSH) and corticotropin‐releasing hormone. Orexin A appeared to be slightly elevated in Fischer rats and cocaine amphetamine‐regulated transcript (CART)55−102 diminished in Brown Norway. At the mRNA level, orexin A, GHSR1, α‐MSH and CART expression were higher in Wistar and Lou C rats. Principal component analysis confirmed the presence of two main factors in the ad libitum rat population; the first being associated with growth‐related parameters and the second being associated with food intake regulation. Hypothalamic GHRH and somatostatin content were positively correlated with feeding‐related neuropeptides such as α‐MSH for GHRH, and orexin A and CART for both peptides. Plasma ghrelin levels were negatively correlated with leptin and IGF‐1 levels. Finally, a 72‐h fasting period affected minimally body weight, plasma IGF‐1 and leptin levels in Lou C rats compared to the three other strains, and plasma insulin levels were less affected in Brown Norway rats. In conclusion, Wistar shorter life span is consistent with its already fatter phenotype at 3 months of age. In terms of IGF‐1, glycaemia and leptin responses to fasting, the Lou strain, which presents with a low food intake/body weight and caloric efficiency, is the least affected. The link between food intake regulation, GH axis and ageing is further demonstrated by principal component analysis, where GHRH and somatostatin were found to be strongly associated with energy homeostasis parameters.
High interdigestive and postprandial motilin levels in patients with the irritable bowel syndrome
Authors: Simrén M. , Björnsson E. S. , Abrahamsson H.
Published: January 2005

Abstract  Motilin shows cyclic variation with the different phases of the migrating motor complex (MMC). Altered motilin levels have been found in irritable bowel syndrome (IBS) patients, but in these studies motilin levels were analysed without the knowledge of the phases of MMC. We included 13 healthy controls (HC) and 24 patients with IBS [12 diarrhoea‐predominant (IBS‐D) and 12 constipation‐predominant (IBS‐C)]. We performed interdigestive and postprandial antroduodenojejunal manometry and blood samples for analysis of motilin were drawn. Group differences in plasma levels of motilin were analysed during mid‐phase II, just before the start of phase III (pre‐III), during phase I, immediately before the meal and 30 and 60 min after the 500 kcal mixed meal. Higher motilin levels were observed in IBS vs HC in both the interdigestive and postprandial periods (P < 0.05). No significant differences between IBS‐C and IBS‐D were observed. The cyclic variation of motilin during MMC and the meal response was similar in IBS and controls. IBS patients, irrespective of the predominant bowel habit, demonstrate higher motilin levels than HCs in all phases of the MMC and also after a meal. These findings may bear some pathophysiological importance in IBS and relate to the gastrointestinal dysmotility often seen in these patients.
Reproducibility of gastric tone, compliance and gastric accommodation assessed with barostat in conscious dogs
Authors: Xing J. H. , Chen J. D. Z.
Published: January 2005

Abstract  The reproducibility of barostat measurements was unclear. In this study, the intraday and interday reproducibility of barostat measurements of gastric tone, compliance and gastric accommodation were assessed in a canine model. A series of experiments were performed using a barostat system in 11 surgically prepared healthy dogs: (i) interday gastric tone and compliance: three sessions on three separate days; (ii) intraday gastric tone and compliance: two sessions on the same day separated by a 30‐min interval; (iii) interday gastric accommodation: two sessions on two separate days, with each including a 30‐min baseline and a 60‐min postprandial period. The results were (i) interday gastric tone (81.2 ± 7.5 mL vs 89.2 ± 8.1 mL vs 86.2 ± 13.6 mL, n = 11) and compliance (n = 8) were comparable; (ii) intraday gastric tone (87.9 ± 17.2 mL vs 77.0 ±14.8 mL, n = 8) and compliance (n = 8) was also similar, but with considerable individual variance; (iii) interday gastric accommodation was 320.8 ±45.1 mL vs 287.9 ± 31.2 mL, no significant difference (n = 8). Inter‐ and intraday gastric tone and compliance and interday gastric accommodation were relatively reproducible in most animals when tested under well‐controlled conditions. However, considerable variations may occur in fasting gastric tone and compliance measurements in certain individuals and cautions should be given when interpreting related results.
Sensitization of mesenteric afferents to chemical and mechanical stimuli following systemic bacterial lipopolysaccharide
Authors: Liu C.‐Y. , Jiang W. , Müller M. H. , Grundy D. , Kreis M. E.
Published: January 2005

Abstract  Background:  The mechanisms underlying endotoxin‐induced hyperalgesia remain unknown. We aimed to study the mechanisms underlying the sensitizing action of lipopolysaccharide (LPS) on intestinal afferent responses to mechanical and chemical stimuli.

Methods:  Extracellular recordings of jejunal afferent nerve discharge were obtained from pentobarbitone‐anaesthetized rats.

Results:  Lipopolysaccharide (6 mg kg−1, i.v.) stimulated a short‐term, transient (<30 min) increase in chemosensitivity to systemic 5‐HT (6 μg kg−1) and responses to mechanical distension and a delayed but maintained (>30 min) increase in spontaneous afferent discharge. Naproxen (10 mg kg−1) and the prostaglandin receptor antagonist AH6809 (1 mg kg−1) significantly attenuated both the short‐term sensitization to mechanical distension and 5‐HT and the long‐term increase in baseline afferent firing following LPS. In contrast, the iNOS inhibitor aminoguanidine (15 mg kg−1) and the L‐type calcium channel antagonist nifedipine (1 mg kg−1) both prolonged the period of afferent sensitization to distension and 5‐HT without influencing the augmented baseline‐firing rate. ω‐Conotoxin GVIA attenuated the increase in afferent discharge to LPS, without any change in mechano‐ and chemosensitivity.

Conclusions:  The long‐term (>30 min) increase in afferent firing following systemic LPS involves neurogenic release of prostanoids. The short‐term (<30 min) sensitization also appears to depend on prostanoid release, while nitric oxide production may serve to down‐regulate LPS‐induced afferent hypersensitivity.
Does the nutrient drink test accurately predict postprandial gastric volume in health and community dyspepsia?
Authors: Gonenne J. , Castillo E. J. , Camilleri M. , Burton D. , Thomforde G. M. , Baxter K. L. , Zinsmeister A. R.
Published: January 2005

Abstract  Nutrient drink tests have been proposed as a surrogate for measurement of gastric accommodation. To study the relationship of maximum tolerated volume (MTV) during nutrient drink test and gastric volumes measured by single‐photon emission computed tomography (SPECT) in healthy controls and functional dyspepsia (FD) patients. We reviewed data from 85 healthy controls and 35 FD residents of south‐eastern Minnesota. All underwent standardized nutrient drink and SPECT studies between August 2000 and June 2003. To test for associations between nutrient drink test and SPECT gastric volumes, we used multiple linear regression and partial regression analyses, assigning age, gender, dyspepsia status and postprandial symptoms as covariates in the model. In the combined group (healthy and FD), MTV was weakly associated with fasting gastric volume (r = 0.43, P = 0.0001) and with volume response to feeding (r = 0.25, P = 0.006). In the FD group, associations were similar (fasting r = 0.53, P = 0.001; postmeal r = 0.32, P = 0.06). After accounting for covariates, MTV only explained 13 and 3% of variations in fasting and postprandial volumes measured by SPECT. MTV during the nutrient drink test does not accurately reflect gastric volume measurements by SPECT in healthy controls and a sample of people in the community with FD.
Contractile response to colonic distention is influenced by oscillation frequency
Authors: Bharucha A. E. , Dhamija S. , Japp A. , Seide B. , Walters B. , Stroetz R. , Zinsmeister A. R. , Hubmayr R. D.
Published: January 2005

Abstract  Static colonic mechanical properties are characterized by stepwise balloon distention. It is unclear whether the state of contractile activation affects frequency‐dependent differences in biomechanical properties. Our aim was to investigate the frequency‐dependence of colonic mechanical properties by sinusoidal oscillation. A descending colonic balloon was sinusoidally oscillated by 25 mL at 5, 10 and 20 cpm in randomized order for 20 min at each frequency in six healthy subjects before and after neostigmine. Volume oscillation was between 75–100 mL before, and 25–50 mL after neostigmine. Pressure waveforms were most variable shortly after commencing oscillation, reflecting an initial contractile response to distention. Elastance (i.e. pressure response to imposed volume) and hysteresivity were estimated; hysteresivity represents the proportion of energy added to the system during inflation, which cannot be recovered during deflation. Colonic elastance was frequency dependent, being highest and most variable at 10 cpm. In contrast, hysteresivity was not significantly different across frequencies. Neostigmine increased mean colonic elastance at all frequencies, and hysteresivity only at 5 cpm. Thus, colonic mechanical properties, particularly elastance are frequency‐dependent. The frequency‐dependence of colonic mechanical properties is worthy of future study because it may provide insights into reflex responses in health and disease.
Motility patterns and distribution of interstitial cells of Cajal and nitrergic neurons in the proximal, mid‐ and distal‐colon of the rat
Authors: Albertí E. , Mikkelsen H. B. , Larsen J. O. , Jiménez M.
Published: January 2005

Abstract  The aim of this work was to study the patterns of spontaneous motility in the circular and longitudinal muscle strips and to characterize the distribution of c‐kit positive interstitial cells of Cajal (ICCs) and nitrergic neurons (nNOS) in the proximal, mid‐ and distal‐colon of Sprague–Dawley rats. We described two types of spontaneous contractions: high frequency (HF) and low frequency (LF) contractions, which were recorded in the presence of tetrodotoxin, suggesting a non‐neurogenic origin. Regional differences were found in the motility patterns depending on the muscle layer and on the part of the colon studied. Muscle strips without submuscular plexus (SMP) showed only LF contractions. The density of ICCs was of the same magnitude along the extent of the colon: about 90–120 cells mm−2 at Auerbach's plexus (AP) and 50–60 cells mm−2 at the SMP. nNOS positive cells were found at the level of the AP and the major density was found in the mid‐colon. Electrical field stimulation abolished LF but did not affect HF contractions. Our results indicate that HF contractions are due to the ICC network found associated with the submuscular plexus (ICC–SMP). The origin of LF contractions is still unknown.
Plasticity of the enteric nervous system during intestinal inflammation
Authors: Lomax A. E. , Fernández E. , Sharkey K. A.
Published: January 2005

Abstract  Inflammation of the bowel causes structural and functional changes to the enteric nervous system (ENS). While morphological alterations to the ENS are evident in some inflammatory conditions, it appears that relatively subtle modifications to the neurophysiology of enteric microcircuits may play a role in gastrointestinal (GI) dysfunction. These include changes to the excitability and synaptic properties of enteric neurones. The response of the ENS to inflammation varies according to the site and type of inflammation, with the functional consequences depending on the nature of the inflammatory stimulus. It has become clear that inflammation at one site can produce changes that occur at remotes sites in the GI tract. Immunohistochemical data from patients with inflammatory bowel disease (IBD) and animal models indicate that inflammation alters the neurochemical content of some functional classes of enteric neurones. A growing body of evidence supports an active role for enteric glia in neuronal and neuroimmune communication in the GI tract, particularly during inflammation. In conclusion, plasticity of the ENS is a feature of intestinal inflammation. Elucidation of the mechanisms whereby inflammation alters enteric neural control of GI functions may lead to novel treatments for IBD.
Obstipation as a paraneoplastic presentation of small cell lung cancer: case report and literature review
Authors: Jun S. , Dimyan M. , Jones K. D. , Ladabaum U.
Published: January 2005

Abstract  Paraneoplastic symptoms caused by abnormal gastrointestinal motility may be the initial manifestation of small cell lung cancer (SCLC). We report a case of a 63‐year‐old woman who presented with progressive constipation culminating in obstipation, and associated symptoms of more widespread dysmotility. A paraneoplastic syndrome was suspected. The only abnormality on chest computed tomography was a minimally enlarged paratracheal lymph node. Positron emission tomography demonstrated increased activity in the lymph node. The antinuclear neuronal antibody titer was elevated. Bronchoscopy with transtracheal biopsy confirmed the diagnosis of SCLC. One year after diagnosis, the patient had progressive symptoms of intestinal obstruction, and ultimately feculent vomiting. On abdominal radiography, colonic sitz markers ingested a year earlier were in virtually the same positions as after ingestion. Palliative colectomy with ileostomy was performed. The myenteric plexus in the terminal ileum and colon showed infiltration by a mixture of B‐cell and T‐cell lymphocytes and plasma cells, and no gross neuronal abnormalities. We review the clinical and pathologic features, clinical course, and management of paraneoplastic pseudoobstruction.
Effects of the selective serotonin reuptake inhibitor, fluoxetine, on regional gastric contractility1
Authors: James A. N. , Ryan J. P. , Parkman H. P.
Published: January 2005

Abstract  Selective serotonin reuptake inhibitors (SSRIs) are increasingly used to treat a variety of disorders but have gastrointestinal side‐effects.

Aim:  To determine the effects of the SSRI, fluoxetine, on gastric smooth muscle contractility.

Methods:  Fundic, antral, and pyloric circular muscle contractility of guinea pig muscle strips were measured in vitro. Fluoxetine was added in concentrations from 0.1 nmol L−1 to 100 μmol L−1. Receptor antagonists were used to determine the neural pathways involved.

Results:  Fluoxetine caused concentration dependent contractions, which were greatest in fundus compared with the antrum or pylorus. The contractile effects of fluoxetine in the antrum were reduced by tetrodotoxin, atropine, phentolamine, and the 5‐HT4 receptor antagonist GR 113808. The contractile effects of fluoxetine in the fundus were reduced by atropine, phentolamine, and GR 113808.

Conclusions:  Fluoxetine affects gastric contractility with regional variability – contracting the fundus more than the antrum or pylorus. The fluoxetine contractile effect is reduced by tetrodotoxin, atropine, phentolamine, and a 5‐HT4 receptor antagonist. These results suggest fluoxetine interacts with muscarinic, alpha‐adrenergic, and serotoninergic receptors and/or ongoing reuptake/release of serotonin in the stomach.
Circumnutation of rice coleoptiles: its occurrence, regulation by phytochrome, and relationship with gravitropism
Authors: YOSHIHARA TAKESHI , IINO MORITOSHI
Published: January 2005

It has been found that coleoptiles of dark‐grown rice (Oryza sativa L.) seedlings undergo regular circumnutation in circular orbits with periods of about 180 min. Both clockwise and counter‐clockwise movements were observed, but individual coleoptiles continued to rotate only in one direction. Light‐grown seedlings did not show circumnutation. In fact, dark‐grown seedlings were found to cease circumnutating in response to a pulse of red light (R). This light‐induced inhibition of circumnutation was demonstrated to involve both a FR‐inducible very‐low‐fluence response, solely mediated by phytochrome A, and a FR‐reversible low‐fluence response, mediated by phytochrome B and/or C. The R‐induced inhibition of circumnutation showed temporal agreement with the R‐induced inhibition of coleoptile growth, suggesting that the former results from the latter. However, about 25% of growth activity remained after R treatment, indicating that circumnutation is more specifically regulated by phytochrome. The R‐treated coleoptile showed gravitropism. Investigation of the growth differential for gravitropic curvature revealed that gravitropic responsiveness was rather enhanced by R. The results suggested that gravitropism is not a cause of circumnutation. It remained probable, however, that gravity perception is a part of the mechanism of circumnutation. It is speculated that the circumnutation investigated aids the seedling shoot in growing through the soil.
OsAGAP, an ARF‐GAP from rice, regulates root development mediated by auxin in Arabidopsis
Authors: ZHUANG X. , XU Y. , CHONG K. , LAN L. , XUE Y. , XU Z.
Published: January 2005

Arf (ADP‐ribosylation factor) proteins, which mediate vesicular transport, have little or no intrinsic GTPase activity. They rely on the action of GTPase‐activating proteins (GAPs) and guanine nucleotide exchange factors (GEFs) for their function. In the present study the OsAGAP gene in rice, which encoded a protein with predicted structure similar to ArfGAP, was identified. The purified OsAGAP‐GST fusion protein was able to stimulate the GTPase activity of rice Arf. Furthermore, OsAGAP can rescue the defect of vesicular transport in the yeast gcs1Δglo3Δ double‐mutant cells. Transgenic Arabidopsis with OsAGAP constitutively expression showed reduced apical dominance, shorter primary roots, increasing number of longer adventitious roots. Many of the phenotypes can be phenocopied by treatment of exogenous indoleacetic acid level (IAA) in wild‐type plants. Determination of whole‐plant IAA level showed that there is a sharp increase of free IAA in OsAGAP transgenic Arabidopsis seedlings. In addition, removal of the 4‐day‐old shoot apex could inhibit the adventitious root formation in the transgenic seedlings. These results suggest OsAGAP, an ARF‐GAP of rice, may be involved in the mediation of plant root development by regulating auxin level.
Within‐stem oxygen concentration and sap flow in four temperate tree species: does long‐lived xylem parenchyma experience hypoxia?
Authors: SPICER R. , HOLBROOK N. M.
Published: January 2005

Oxygen levels as low as 1–5% (gaseous mole fraction) occur in secondary xylem, but it is not known if there is a consistent pattern of decline in O2 from the cambium toward the pith, or whether parenchyma cells experience hypoxic conditions deep within the stem. We developed a system for repeated in situ measurement of O2 at different depths within stems of Acer rubrum, Fraxinus americana, Tsuga canadensis, and Quercus rubra. In summer during active transpiration, O2 declined from the cambium toward the heartwood boundary in F. americana, T. canadensis and Q. rubra, but remained constant in A. rubrum. Average sapwood O2 was about 10%, with the lowest values observed in the innermost sapwood around 3–5%. Before spring leaf flush, O2 content in the outer sapwood was reduced in Q. rubra and T. canadensis relative to summer, and was occasionally lower than in the inner sapwood. Sapwood respiration in T. canadensis was constant above 5% O2, but reduced by about 65% at 1% O2. In F. americana, sapwood respiration was constant above 10% O2 but reduced by 25% at 5% O2, and by 75% at 1% O2, the most extreme inhibition observed. However, when prolonged (72 h) exposure to 1%, 5% and 10% O2 was followed by re‐equilibration to 10% O2, no inhibition was found. Given the minor (and reversible) effect of low O2 on parenchyma metabolism at levels common in the inner sapwood, it is unlikely that O2 content severely limits parenchyma respiration or leads to parenchyma cell death during sapwood senescence. Within‐stem O2 levels may instead be most relevant to metabolism in the cambial zone and phloem, for which sapwood could serve as a significant source of O2.
Salt‐stress signalling and the role of calcium in the regulation of the Arabidopsis ATHB7 gene
Authors: HENRIKSSON E. , NORDIN HENRIKSSON K.
Published: January 2005

In plants changes in cytosolic calcium ion concentrations ([Ca2+]cyt) have been detected after various stress treatments, including salt treatment. The involvement of a Ca2+ signal as an essential component of signalling pathways leading to downstream responses, such as gene expression, is supported only by a few studies. In this study the possible involvement of the salt stress‐induced increase in [Ca2+]cyt in the signalling pathway leading to the induction of ATHB7, a homeobox gene encoding a homeodomain leucine zipper (HDZip) transcription factor was analysed. The salt‐induced expression of ATHB7 was found to be independent of the Ca2+ signal evoked by salt. Instead, it was found that ATHB7 expression in shoots was not dependent on a direct contact with salt or osmoticum, whereas in roots, ATHB7 seemed to be induced by the direct contact, indicating that signals from roots cause systemic induction of ATHB7. Abscisic acid (ABA) or ABA‐dependent components were found to, at least partly, to function as the systemic signal.