Metallothionein-3 (MT-3), also known as growth inhibitory factor (GIF), was originally identified in the brain. An essential step in elucidating the potential roles of MT-3 is to evaluate its expression levels in organs other than the brain. In this present study, we carried out RT-PCR, Western blot and immunohistochemical analyses to quantify MT-3 mRNA and its protein in the cerebrum, eye, heart, kidney, liver, prostate, testis, tongue, and muscle in male Wistar rats. MT-3 mRNA was detected in the cerebrum, the dorsolateral lobe of the prostate, testis, and tongue. Using a monoclonal anti-MT-3 antibody, we detected MT-3 in the cerebrum, the dorsolateral lobe of the prostate, testis, and tongue as a single band on an immunoblot. Immunohistochemical staining showed MT-3 in some astrocytes in the deep cortex, ependymal cells, and choroidal cells in the cerebrum. MT-3 was also detected in some cells of the glomerulus and the collective tubules in the kidney, some cells in the glandular epithelium of the dorsolateral lobe of the prostate, some Sertoli cells and Lydig cells in the testis, and taste bud cells in the tongue. Although MT-3 immunopositivity was obviously demonstrated in the kidney by the immnunohistochemical method, the expression of MT-3 was not fully detectable by RT-PCR and Western blot analyses. Interestingly, only a subset of cells showed positivity for MT-3, not all cells in all tissues. The localization of MT-3 in peripheral organs outside the brain suggests that MT-3 has roles in these tissues besides its role in growth inhibition of neurites.

OBJECTIVE

Each year at our institution we have been performing over 100 endoscopic mucosal resections (EMRs) for gastric tumor. However, there are some tumor locations where it is difficult to carry out this procedure, such as the lesser curvature or posterior wall of the gastric body, the cardia, and the lesser curvature of the antrum. To facilitate EMR of tumors in these locations, a multibending scope (the "M-scope", Olympus GIF-2T240M; Olympus Optical, Tokyo, Japan) has been developed, which has two independently curving segments. The aim of this study was to evaluate the effectiveness of this new multibending scope for EMR of gastric tumors.

METHODS

Using the M-scope, we carried out EMR in 59 patients at the Jikei University Hospital. The lesions were located in the cardia in seven patients, in the gastric body in 30 patients, and in the antrum in 22 patients. The effectiveness of the M-scope was evaluated by experienced endoscopists; a score of 1 was given for very good effectiveness, and a score of 0 was given when there was no notable difference in effectiveness from the conventional scope. Evaluation was done according to the location of the lesions, the method used for EMR, and the tumor diameter. When the score for each item was equivalent to the average overall score for all the procedures or higher, the M-scope was defined as being effective.

RESULTS

The overall score for all the procedures was 0.7 +/- 0.4 (average +/- SD) (very good in 43 procedures; no notable difference in 16 procedures). When assessed according to the location of the lesion, the mean effectiveness scores of the M-scope for lesions at the following locations were higher than the average overall score: the lesser curvature of the antrum (0.8 +/- 0.5); the posterior wall of the gastric body (1.0 +/- 0.0); the greater curvature of the gastric body (1.0 +/- 0.0); and the lesser curvature of the gastric body (0.9 +/- 0.4). The results suggested that the M-scope was effective for EMR of gastric tumors at these traditionally difficult locations. With regard to the scores assessed according to the method of EMR, the mean scores were 0.8 +/- 0.4 for the two-channel scope method and 0.9 +/- 0.4 for EMR using an insulated-tip diathermic knife (IT-EMR), again suggesting that the M-scope was effective for EMR by these methods. When evaluated according to tumor size, the score was 0.8 +/- 0.4 when the tumor was 11 mm or greater in diameter, indicating that the M-scope was effective for EMR of large tumors.

CONCLUSIONS

The results of the study suggest that the M-scope is effective for EMR of tumors in the lesser curvature of the antrum, and in the posterior wall, lesser curvature, or greater curvature of the gastric body. With regard to the method of EMR, the M-scope is effective for both the two-channel scope method and IT-EMR. In relation to tumor size, the M-scope is effective for the resection of large tumors.

One hundred and forty-two pediatric patients between age 1 month and 20 years had 163 endoscopic procedures. Of 66 with chronic abdominal pain, 21 had a source identified endoscopically that was seen in only 15 by esophagogram and upper gastrointestinal series. Of 31 with nausea, vomiting, dysphagia, and/or odynophagia and retrosternal pain, endoscopy demonstrated the source in 19 patients and radiographic studies in 14. Of 34 with hematemesis and/or melena, 26 had a bleeding site identified endoscopically but only 4 of 28 had an identified source by radiographic studies. Duodenal and gastric ulcers and hemorrhagic gastritis were the commonest cases of upper gastrointestinal bleeding and organically of chronic adbominal pain. Functional abdominal pain was the commonest cause of chronic abdominal pain in those endoscoped. Foreign bodies were removed from the esophagus and stomach of 6 patients and dislodged in 2 others. Caustic ingestion was recognized in the esophagus and stomach of 2 patients who did not have mouth burns. The GIF-P2-prototype with four-way tip control and ability to retroflex 180 degree up, 60 degree down, and 100 degree right and left was superior to GIF-P1 and CF-P-prototype for visualization of the entire esophagus, stomach, duodenal bulb, and postbulbar area in patients less than 10 years old. Visualization of the duodenal bulb was possible in 28 of 29 pediatric patients, and of the postbulbar area in 25 of 26 in whom it was attempted. Infants who weighed as little as 3 to 5 kg were successfully examined. Retroflexion was possible in 29 of 30 to see the fundus and cardioesophageal junction. Patients older than 10 years were better examined with the GIF-D because of its increased ability to transmit light. Sedation for the school-age child with 0.5 to 1.0 mg per kg of diazepam and 1 to 2 mg per kg of meperidine given intravenously provides excellent sedation in most instances. General anesthesia is preferable for the preschooler and infant. Minor complications occurred in 2 patients who received less than adequate sedation and in 1 patient with general anesthesia.

BACKGROUND

Heterogeneity of databases and software resources continues to hamper the integration of biological information. Top-down solutions are not feasible for the full-scale problem of integration across biological species and data types. Bottom-up solutions so far have not integrated, in a maximally flexible way, dynamic and interactive graphical-user-interface components with data repositories and analysis tools.

RESULTS

We present a component-based approach that relies on a generalized platform for component integration. The platform enables independently-developed components to synchronize their behavior and exchange services, without direct knowledge of one another. An interface-based data model allows the exchange of information with minimal component interdependency. From these interactions an integrated system results, which we call ISYSf1.gif" BORDER="0">. By allowing services to be discovered dynamically based on selected objects, ISYS encourages a kind of exploratory navigation that we believe to be well-suited for applications in genomic research.

A catheter-type endomicroscope has been developed with a maximum magnifying power of 1100 times. Living cancer cells in the esophagus, stomach, and colon were successfully observed in high-resolution images. The "Endo-Cytoscopy system" (prototype,Olympus Optical, Co., Tokyo, Japan) is a catheter-based probe capable of passage through the accessory channel of the endoscope(GIF-1T, Olympus). Methylene blue solution was used for vital staining of the in vivo gastrointestinal mucosa. Living cells in both normal mucosa and malignant tissue were clearly demonstrated in luminal organs. In particular, the nucleus, cell body, and nucleolus were clearly demonstrated with high-quality images similar to those of conventional cytology. This novel technology has the potential to provide an in vivo histologic diagnosis via "optical biopsy" and virtual histology.

The subcellular localization of growth inhibitory factor (GIF), a brain-specific member of the metallothionein family, was determined in the rat brain by electron microscopic immunohistochemistry using a rabbit antiserum against a synthetic polypeptide specific for rat GIF. The major cell type that expressed a high level of GIF immunoreactivity was the astrocytes. In these cells, dense labelling was observed throughout the soma and the fine processes, in association with the free ribosomes, rough endoplasmic reticulum, small vesicles, the outer membrane of the mitochondria and part of the plasma membrane. Astrocytic end-feet around blood vessels exhibited intense immunoreactivity. Another cell type exhibiting GIF immunolabelling was the neurons. However, this immunoreactivity was restricted to a subset of the neuronal population, and in contrast to the astrocytic pattern, the labelling was localized predominantly in the processes including axons and dendrites, in association with microtubules, ribosomes, the outer membrane of the mitochondria and the plasmalemma. Synaptic elements, including dendritic spines, also showed definite immunoreactivity in association with synaptic vesicles and post-synaptic densities. No labelling was observed in the oligodendrocytes or microglia. The present data suggest that GIF is expressed in both astrocytes and neurons, and plays rather specific roles in each phenotype.

Metallothionein-III is a low molecular weight, heavy-metal binding protein expressed mainly in the central nervous system. First identified as a growth inhibitory factor (GIF) of rat cortical neurons in vitro, it has subsequently been shown to be a member of the metallothionein (MT) gene family and renamed as MT-III. In this study we have raised polyclonal antibodies in rabbits against recombinant rat MT-III (rMT-III). The sera obtained reacted specifically against recombinant zinc-and cadmium-saturated rMT-III, and did not cross-react with native rat MT-I and MT-II purified from the liver of zinc injected rats. The specificity of the antibody was also demonstrated in immunocytochemical studies by the elimination of the immunostaining by preincubation of the antibody with brain (but not liver) extracts, and by the results obtained in MT-III null mice. The antibody was used to characterize the putative differences between the rat brain MT isoforms, namely MT-I+II and MT-III, in the freeze lesion model of brain damage, and for developing an ELISA for MT-III suitable for brain samples. In the normal rat brain, MT-III was mostly present primarily in astrocytes. However, lectin staining indicated that MT-III immunoreactivity was also present in microglia, monocytes and/or macrophages in the leptomeninges and lying adjacent to major vessels. In freeze lesioned rats, both MT-I+II and MT-III immunoreactivities increased in the ipsilateral cortex. The pattern of MT-III immunoreactivity significantly differed from that of MT-I+II, since the latter was evident in both the vicinity of the lesioned tissue and deeper cortical layers, whereas that of the former was located only in the deeper cortical layers. This suggests different roles for these MT isoforms, and indeed in a new bioassay measuring astrocyte migration in vitro, rMT-III promoted migration to a higher extent than MT-I+II. Thus, MT-III could not only affect neuronal sprouting as previously suggested, but also astrocyte function. Finally, MT-III protein levels of patients with Alzheimer's disease (AD) were, if anything, increased when compared with similarly aged control brains, which was in agreement with the significantly increased MT-III mRNA levels of AD brains.

Contradictory results have been reported on the downregulation and role of the brain-specific protein metallothionein-III (MT-III, GIF) in Alzheimer disease (AD). In this article, the importance of MT-III downregulation in AD brain was re-evaluated in temporal and frontal cortex, hippocampus, and cerebellum of 11 AD patients and two groups of five and six control subjects, respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was used to quantify the levels of MT-III mRNA relative to the levels of three constitutive RNAs: beta-actin, glyceraldehyde-3-phosphate dehydrogenase (G3PDH), and ribosomal RNA 18S (rRNA 18S). The distribution of MT-III was similar to that of each of the three constitutive RNAs. The relative levels of each of these RNAs was high in brain regions examined in both AD patients and control subjects. Our findings do not support a downregulation of MT-III mRNA in the frontal cortex as well as the temporal cortex and hippocampus of AD patients. However, the level of MT-III mRNA was not constant in the investigated samples, suggesting that MT-III mRNA regulation could be controlled by factors other than AD pathology. Brain-derived neurotrophic factor (BDNF) mRNA levels were hardly detectable by RT-PCR in human brain tissue; a trend for a decrease was apparent in the temporal cortex of AD patients. In conclusion, the content of MT-III mRNA in the brain of AD patients was not detectably impaired, whereas BDNF mRNA may be affected.

BACKGROUND

A genetically influenced atherogenic lipoprotein phenotype characterized by a predominance of small, dense LDL particles (subclass pattern B) can be induced by low-fat diets in healthy subjects with large LDL particles (pattern A).

OBJECTIVE

The aim of this study was to test whether genetic predisposition to subclass pattern B contributes to susceptibility to induction of this trait by a low-fat diet.

METHODS

The prevalence of pattern B in children is relatively low compared with that in older individuals, but genetic susceptibility to this trait in offspring can be inferred by its presence in their parents. Plasma lipoproteins were analyzed 10 d after a change from a usual diet to a very-low-fat (10% fat), high-carbohydrate diet in offspring (mean age: 14 y; range: 7-28 y) of 22 families according to parental LDL-subclass patterns when consuming a low-fat diet: AxA mating (9 families with 19 children), AxB mating (5 families with 10 children), and BxB mating (8 families with 21 children).

RESULTS

The very-low-fat, high-carbohydrate diet produced significantly greater decreases in LDL particle size in offspring of BxB parents (f1.gif" BORDER="0"> +/- SE: -0.55 +/- 0.16 nm) and AxB parents (-0.48 +/- 0.19 nm) than in offspring of AxA parents (0.14 +/- 0.20 nm). The number of children expressing pattern B with the 10%-fat diet and the proportion of children converting from pattern A to pattern B was significantly greater in offspring of BxB parents than in those with 1 or 2 pattern A parents.

CONCLUSIONS

A very-low-fat, high-carbohydrate diet can induce expression of LDL-subclass pattern B in genetically predisposed children with low expression of the trait while consuming their usual diets.

CONCLUSIONS

Several programs are now available for analyzing the large datasets arising from cDNA microarray experiments. Most programs are expensive commercial packages or require expensive third party software. Some are freely available to academic researchers, but are limited to one operating system. MicroArray Genome Imaging and Clustering Tool (MAGIC Tool) is an open source program that works on all major platforms, and takes users 'from tiff to gif'. Several unique features of MAGIC Tool are particularly useful for research and teaching.

BACKGROUND

http://www.bio.davidson.edu/MAGIC

Attempts were made to generate Ag-specific suppressor T cells from Ag-primed spleen cells by using glycosylation inhibiting factor (GIF). BDF1 mice were primed with alum-absorbed OVA and their spleen cells were stimulated with OVA. Ag-activated T cells were then propagated in IL-2-containing conditioned medium. Incubation of the T cells with OVA-pulsed syngeneic macrophages resulted in the formation of IgE-potentiating factor and glycosylation-enhancing factor that has affinity for OVA, i.e., OVA-specific glycosylation-enhancing factor. However, if the same Ag-activated splenic T cells were propagated in the IL-2-containing medium in the presence of GIF T cells obtained in the cultures formed IgE-suppressive factors and OVA-specific GIF on antigenic stimulation. Thus we constructed T cell hybridomas from the Ag-activated T cells propagated by IL-2 in the presence of GIF. A representative hybridoma, 71B4, formed OVA-specific GIF on incubation with OVA-pulsed macrophages of BDF1 mice or C57B1/6 mice. However, if the same hybridoma cells were incubated with OVA alone or with OVA-pulsed macrophages of H-2k or H-2d strains, they produced GIF that had no affinity for OVA. The OVA-specific GIF bound to OVA-Sepharose but did not bind to BSA-Sepharose or KLH Sepharose. Intravenous injections of the OVA-specific GIF from the hybridoma suppressed the IgE and IgG1 anti-DNP antibody response of BDF1 mice to DNP-OVA, but failed to suppress the anti-hapten antibody responses of the strain to DNP-keyhole limpet hemocyanin, indicating that the factors suppressed the antibody response in a carrier-specific manner. However, the same OVA-specific GIF failed to suppress the anti-hapten antibody response of DBA/1 mice to DNP-OVA, suggesting that the immunosuppressive effects of the factors is MHC restricted.

The first gamma-2 (gamma2) heavy chain disease protein Gif has pyrroli-dinecarboxylic acid as its amino terminal residue, much of the Fd variable region, and an internal deletion of the heavy chain of about 100 residues corresponding to most of the Fd constant region. Normal sequence resumes with a glutamic acid residue at position 216 in the hinge region. This is the third gamma heavy chain disease protein where normal sequence resumes at the same position after the deletion.

Growth-inhibitory factor (GIF), which is deficient in the Alzheimer's disease brain, is a 68-amino-acid metallothionein-like protein. GIF is expressed in the central nervous system but not in peripheral tissues including dorsal root ganglion. GIF is immunocytochemically distributed in Bergmann's glia in the cerebellum and astrocytes in the neocortex, hippocampus, striatum, brain stem, and spinal cord. Strong GIF immunoreactivity is localized in astrocyte cell bodies and fine long processes, which are closely associated with neural perikarya and dendrites, such as layers 2-6 of cerebral gray matter, the molecular layer of the dentate gyrus, the pyramidal layer of the hippocampus and spinal gray matter. GIF is induced in reactive astrocytes in the cerebral cortex in cases of meningitis and Creutzfeldt-Jakob disease or in reactive astrocytes surrounding old cerebral infarcts. On the other hand, GIF is reduced in the subset of reactive astrocytes in lesioned areas of degenerative diseases such as Alzheimer's disease, multiple-system atrophy, Parkinson's disease, progressive supranuclear palsy and amyotrophic lateral sclerosis. Reduction of GIF is correlated with neuronal loss. Thus, perturbation in normal neuroglial interaction in degenerative diseases may lead to a reduction of GIF in reactive astrocytes.

Five different Ag-binding suppressor factors from two types of hapten-specific Ts cell hybridomas (TsF1 inducer and TsF3 effector factors) were bound by an anti-lipomodulin mAb (141-B9), that crossreacts with rodent glycosylation inhibition factor (GIF). The Ag-specific suppressor activity in these hybridoma supernatants was bound by anti-lipomodulin columns and could be recovered by elution at acid pH. Additional evidence for the expression of lipomodulin/GIF activity on these TsF molecules was demonstrated by the ability of the eluted fractions to inhibit the glycosylation of IgE-binding peptides during their biosynthesis. The same biologic activity is associated with GIF and lipomodulin. The relationship between TsF and lipomodulin/GIF was confirmed in a serologic assay, which showed that TsF1 and TsF3 molecules, whether purified over Ag, anti-IJ or anti-TsF columns, are recognized by the mAb. 141-B9. The combined results indicate that Ag-binding Ts factors share a common antigenic determinant with phospholipase inhibitory proteins such as lipomodulin and GIF. In addition, the demonstration of glycosylation regulatory activity carried on these TsF molecules suggests a possible mode for their bioactivity.

Previous studies in our laboratory demonstrated an altered immuno-endocrine feedback communication via the hypothalamo-pituitary-adrenal (HPA) axis, which may be an important modulatory factor in the development of spontaneous autoimmune thyroiditis in Obese strain (OS) chickens. These birds show a significantly lower, or even absent, increase in serum glucocorticoid levels in response to an intravenous injection of antigen or conditioned medium (CM) from mitogen-stimulated spleen cells known to contain glucocorticoid-increasing factors (GIFs), notably interleukin-1 (IL-1). The present study was aimed at investigating this feedback regulation in animal models with spontaneous systemic autoimmune diseases, such as the UCD-200 chicken, which serves as a model for human scleroderma, and various murine lupus models. In contrast to OS chickens, UCD-200 chickens displayed a nearly normal plasma corticosterone surge in response to CM, and IL-1 was again identified as the primary GIF in CM. Recombinant IL-1 also induced a drastic increase in plasma corticosterone levels in various strains of normal mice. A similar increase was observed in the bacterial lipopolysaccharide-resistant C3H/HeJ strain, thus excluding the possibility of bacterial endotoxin contamination. However, in young lupus-prone (NZB/W)F1 and MRL/MP-lpr mice, a significantly lower increase in plasma corticosterone levels was observed after injection of recombinant IL-1, suggesting a deficient immuno-endocrine communication via the HPA loop in this instance as well. Detailed studies to identify further cytokines with GIF activity in the avian and murine systems showed that both IL-6 and tumor necrosis factor-alpha could induce increased plasma corticosterone levels in mice, but not in chickens. IL-3, IL-8, transforming growth factor-beta, interferon-gamma and granulocyte-macrophage colony-stimulating factor were devoid of GIF activity in both chickens and mice.

MAnGO (Microarray Analysis at the Gif/Orsay platform) is an interactive R-based tool for the analysis of two-colour microarray experiments. It is a compilation of various methods, which allows the user (1) to control data quality by detecting biases with a large number of visual representations, (2) to pre-process data (filtering and normalization) and (3) to carry out differential analyses. MAnGO is not only a 'turn-key' tool, oriented towards biologists but also a flexible and adaptable R script oriented towards bioinformaticians.

BACKGROUND

http://bioinfome.cgm.cnrs-gif.fr/.

It has long been thought that growth-regulating factors (GRFs) gene family members act as transcriptional activators to play important roles in multiple plant developmental processes. However, the recent characterization of Arabidopsis GRF7 showed that it functions as a transcriptional repressor of osmotic stress-responsive genes. This highlights the complex and diverse mechanisms by which different GRF members use to take action. In this study, the maize (Zea mays L.) GRF10 was functionally characterized to improve this concept. The deduced ZmGRF10 protein retains the N-terminal QLQ and WRC domains, the characteristic regions as protein-interacting and DNA-binding domains, respectively. However, it lacks nearly the entire C-terminal domain, the regions executing transactivation activity. Consistently, ZmGRF10 protein maintains the ability to interact with GRF-interacting factors (GIFs) proteins, but lacks transactivation activity. Overexpression of ZmGRF10 in maize led to a reduction in leaf size and plant height through decreasing cell proliferation, whereas the yield-related traits were not affected. Transcriptome analysis revealed that multiple biological pathways were affected by ZmGRF10 overexpression, including a few transcriptional regulatory genes, which have been demonstrated to have important roles in controlling plant growth and development. We propose that ZmGRF10 aids in fine-tuning the homeostasis of the GRF-GIF complex in the regulation of cell proliferation.

A cDNA sequence coding for rat interleukin-1 alpha (IL-1 alpha) has been isolated from a cDNA library that was prepared with mRNA derived from LPS-stimulated rat peritoneal macrophages by using human IL-1 alpha cDNA as a probe. The rat cDNA encodes a 270 amino acid residue protein which is homologous (65%) to human IL-1 alpha. The rat cDNA sequence under SV40 early promoter directed the synthesis of biologically active IL-1 in monkey COS-1 cells. Rat IL-1 alpha mRNA is not expressed in spleen, lung, liver or brain, and is also not expressed in these organs of LPS-treated rat except spleen. This suggests that IL-1 alpha is not produced constitutively in various tissues and LPS is not sufficient to induce IL-1 alpha in most tissues. Our data indicate that the IL-1 activities which have been reported to be produced in the brain are not of alpha type. We have constructed a plasmid expressing the carboxy terminal 156 amino acids in Escherichia coli. Recombinant rat IL-1 alpha produced in COS cells or E. coli has cytotoxic activity against the human melanoma cell line A375S1 (GIF activity), which has been reported to be sensitive to human IL-1 alpha and IL-1 beta. This suggests that GIF activity is common to IL-1s derived from various sources.

BACKGROUND

Inherited malabsorption of cobalamin (Cbl) causes hematological and neurological abnormalities that can be fatal. Three genes have been implicated in Cbl malabsorption; yet, only about 10% of ~400-500 reported cases have been molecularly studied to date. Recessive mutations in CUBN or AMN cause Imerslund-Gräsbeck Syndrome (IGS), while recessive mutations in GIF cause Intrinsic Factor Deficiency (IFD). IGS and IFD differ in that IGS usually presents with proteinuria, which is not observed in IFD. The genetic heterogeneity and numerous differential diagnoses make clinical assessment difficult.

METHODS

We present a large genetic screening study of 154 families or patients with suspected hereditary Cbl malabsorption. Patients and their families have been accrued over a period spanning >12 years. Systematic genetic testing of the three genes CUBN, AMN, and GIF was accomplished using a combination of single strand conformation polymorphism and DNA and RNA sequencing. In addition, six genes that were contenders for a role in inherited Cbl malabsorption were studied in a subset of these patients.

RESULTS

Our results revealed population-specific mutations, mutational hotspots, and functionally distinct regions in the three causal genes. We identified mutations in 126/154 unrelated cases (82%). Fifty-three of 126 cases (42%) were mutated in CUBN, 45/126 (36%) were mutated in AMN, and 28/126 (22%) had mutations in GIF. We found 26 undescribed mutations in CUBN, 19 in AMN, and 7 in GIF for a total of 52 novel defects described herein. We excluded six other candidate genes as culprits and concluded that additional genes might be involved.

CONCLUSIONS

Cbl malabsorption is found worldwide and genetically complex. However, our results indicate that population-specific founder mutations are quite common. Consequently, targeted genetic testing has become feasible if ethnic ancestry is considered. These results will facilitate clinical and molecular genetic testing of Cbl malabsorption. Early diagnosis improves the lifelong care required by these patients and prevents potential neurological long-term complications. This study provides the first comprehensive overview of the genetics that underlies the inherited Cbl malabsorption phenotype.

Human neuronal growth inhibitory factor (GIF), a metallothionein-like protein classified as metallothionein-3, impairs the survival and the neurite formation of cultured neurons. Despite its approximately 70% amino acid sequence identity with those of mammalian metallothioneins (MT-1 and MT-2 isoforms), only GIF exhibits growth inhibitory activity. In this study, structural features of the metal-thiolate clusters in recombinant Zn(7)- and Cd(7)-GIF, and in part also in synthetic GIF (68 amino acids), were investigated by using circular dichroism (CD) and (113)Cd NMR. The CD and (113)Cd NMR studies of recombinant Me(7)-GIF confirmed the existence of distinct Me(4)S(11)- and Me(3)S(9)-clusters located in the alpha- and beta-domains of the protein, respectively. Moreover, a mutual structural stabilization of both domains was demonstrated. The (113)Cd NMR studies of recombinant (113)Cd(7)-GIF were conducted at different magnetic fields (66.66 and 133.33 MHz) and temperatures (298 and 323 K). At 298 K the spectra revealed seven (113)Cd signals at 676, 664, 651, 644, 624, 622, and 595 ppm. A striking feature of all resonances is the absence of resolved homonuclear [(113)Cd-(113)Cd] couplings and large apparent line widths (between 140 and 350 Hz), which account for the absence of cross-peaks in [(113)Cd, (113)Cd] COSY. On the basis of a close correspondence in chemical shift positions of the (113)Cd signals at 676, 624, 622, and 595 ppm with those obtained in our previous studies of (113)Cd(4)-GIF(32-68) [Hasler, D. W., Faller, P., and Vasák, M. (1998) Biochemistry 37, 14966], these resonances can be assigned to a Cd(4)S(11)-cluster in the alpha-domain of (113)Cd(7)-GIF. Consequently, the remaining three (113)Cd signals at 664, 651, and 644 ppm originate from a Me(3)S(9) cluster in the beta-domain. However, the latter resonances show a markedly reduced and temperature-independent intensity (approximately 20%) when compared with those of the alpha-domain, indicating that the majority of the signal intensity remained undetected. To account for the observed NMR features of (113)Cd(7)-GIF, we suggest that dynamic processes acting on two different NMR time scales are present: (i) fast exchange processes among conformational cluster substates giving rise to broad, weight-averaged resonances and (ii) additional very slow exchange processes within the beta-domain associated with the formation of configurational cluster substates. The implications of the structure fluctuation for the biological activity of GIF are discussed.

FTG is a web server for analyzing nucleotide sequences to predict the genes using Fourier transform techniques. This server implements the existing Fourier transform algorithms for gene prediction and allows the rapid visualization of analysis by output in GIF format.

The authors estimated the generalized impact fraction (GIF) for heart failure (HF) related to obesity, representing the proportion of incident HF events that could be prevented from reductions in obesity and/or overweight. The Atherosclerosis Risk in Communities Study is a biracial population-based cohort study of persons aged 45-64 years from 4 US communities with a median 14 years of follow-up (1987-2003) for incident, hospitalized, or fatal HF. Body mass index (BMI; weight (kg)/height (m)(2)) was measured at baseline (1987-1989) and categorized as normal weight (BMI <25), overweight (BMI 25-29.9), or obese (BMI ≥30). After exclusion of prevalent HF, missing BMI, and poorly represented racial groups, the sample size was 14,642. The GIF and attributable fraction were calculated using a case-load weighted-sum method. A 95% distribution of the GIF was estimated from bootstrapped data sets. A 30% hypothetical reduction in obesity/overweight would potentially prevent 8.5% (95% simulation interval: 6.1, 10.7) of incident HF events. The attributable fraction, which assumes complete elimination of obesity/overweight, was 28% (95% simulation interval: 20, 36)-approximately 3 times larger than the most optimistic GIF calculated here. Investigators studying exposures that are unlikely to be eradicated given current prevention efforts, such as obesity, should consider estimating the GIF to avoid overestimates of population impact.

Event-driven simulation strategies were proposed recently to simulate integrate-and-fire (IF) type neuronal models. These strategies can lead to computationally efficient algorithms for simulating large-scale networks of neurons; most important, such approaches are more precise than traditional clock-driven numerical integration approaches because the timing of spikes is treated exactly. The drawback of such event-driven methods is that in order to be efficient, the membrane equations must be solvable analytically, or at least provide simple analytic approximations for the state variables describing the system. This requirement prevents, in general, the use of conductance-based synaptic interactions within the framework of event-driven simulations and, thus, the investigation of network paradigms where synaptic conductances are important. We propose here a number of extensions of the classical leaky IF neuron model involving approximations of the membrane equation with conductance-based synaptic current, which lead to simple analytic expressions for the membrane state, and therefore can be used in the event-driven framework. These conductance-based IF (gIF) models are compared to commonly used models, such as the leaky IF model or biophysical models in which conductances are explicitly integrated. All models are compared with respect to various spiking response properties in the presence of synaptic activity, such as the spontaneous discharge statistics, the temporal precision in resolving synaptic inputs, and gain modulation under in vivo-like synaptic bombardment. Being based on the passive membrane equation with fixed-threshold spike generation, the proposed gIF models are situated in between leaky IF and biophysical models but are much closer to the latter with respect to their dynamic behavior and response characteristics, while still being nearly as computationally efficient as simple IF neuron models. gIF models should therefore provide a useful tool for efficient and precise simulation of large-scale neuronal networks with realistic, conductance-based synaptic interactions.

Glycosylation inhibiting factor (GIF) was purified from culture filtrates of a T cell hybridoma, 23A4, by affinity chromatography on anti-lipomodulin Sepharose. The factor exhibited phospholipase inhibitory activity upon dephosphorylation. Immunization of BDF1 mice with aluminum hydroxide gel (alum)-absorbed dinitrophenyl derivatives of ovalbumin (DNP-OA) resulted in persistent IgE and IgG antibody formation. However, repeated injections of the affinity-purified GIF into the DNP-OA-primed mice beginning on the day of priming prevented the primary anti-hapten antibody responses of both the IgE and the IgG1 isotypes. Treatment with GIF also diminished on-going IgE antibody formation in the DNP-OA-primed mice. The treatment changed the nature of IgE-binding factors formed by BDF1 spleen cells. Incubation of spleen cells from OA + alum-primed mice with OA resulted in the formation of IgE-potentiating factor, whereas spleen cells of OA-primed, GIF-treated mice formed IgE-suppressive factor upon antigenic stimulation. It was also found that Lyt-2+ T cells in the OA-primed, GIF-treated mouse spleen cells released GIF, which had affinity for OA and bore I-Jb determinant(s). Transfer of a Lyt-1+ cell-depleted fraction of the OA-primed, GIF-treated mouse spleen cells into naive syngeneic animals resulted in suppression of the primary anti-DNP IgE antibody response of the recipients to alum-absorbed DNP-OA, but failed to affect the anti-DNP antibody response to DNP-keyhole limpet hemocyanin. The results indicate that GIF treatment during the primary response to OA facilitated the generation of antigen-specific suppressor T cells.