Glial cell line-derived neurotrophic factor-dependent RET activation can be mediated by two different cell-surface accessory proteins

Glial cell line-derived neurotrophic factor (GDNF)-dependent activation of the tyrosine kinase receptor RET is necessary for kidney and enteric neuron development, and mutations in RET are associated with human diseases. Activation of RET by GDNF has been shown to require an accessory component, GDNFR-alpha (RETL1). We report the isolation and characterization of rat and human cDNAs for a novel cell-surface associated accessory protein, RETL2, that shares 49% identity with RETL1. Both RETL1 and RETL2 can mediate GDNF dependent phosphorylation of RET, but they exhibit different patterns of expression in fetal and adult tissues. The most striking differences in expression observed were in the adult central and peripheral nervous systems. In addition, the mechanisms by which the two accessory proteins facilitate the activation of RET by GDNF are quite distinct. In vitro binding experiments with soluble forms of RET, RETL1 and RETL2 demonstrate that while RETL1 binds GDNF tightly to form a membrane-associated complex which can then interact with RET, RETL2 only forms a high affinity complex with GDNF in the presence of RET. This strong RET dependence of the binding of RETL2 to GDNF was confirmed by FACS analysis on RETL1 and RETL2 expressing cells. Together with the recent discovery of a GDNF related protein, neurturin, these data raise the possibility that RETL1 and RETL2 have distinctive roles during development and in the nervous system of the adult. RETL1 and RETL2 represent new candidate susceptibility genes and/or modifier loci for RET-associated diseases.     

Mutations of p53 gene can be detected in the plasma of patients with large bowel carcinoma

A 55-year-old woman was admitted because of progressive jaundice. Blood examination on admission revealed markedly elevated serum levels of CA19-9 and SPAN-1. Abdominal computed tomography revealed a large tumor in the head of the pancreas. Although the patient's jaundice and elevated CA19-9 decreased after percutaneous franshepatic cholangio-drainage, her SPAN-1 level remained elevated. Open biopsy of the pancreatic tumor revealed non-Hodgkin's lymphoma (NHL) (diffuse medium, B cell type), Complete remission was obtained after one course of CHOP therapy. This case suggests that pancreatic tumor with elevated serum CA19-9 and SPAN-1 levels may involve NHL, and may be curable with chemotherapy.     

Calciuric effects of protein and potassium bicarbonate but not of sodium chloride or phosphate can be detected acutely in adult women and men

An acute load test was used to test the influence of dietary factors on urinary calcium excretion. In study 1, 10 fasting premenopausal women consumed test meals providing a moderate amount of protein (MP; 23 g), MP plus 23 mmol KHCO3 (MP+K), MP plus 23 mmol NaCl (MP+Na), and a high amount of protein (HP; 53 g), HP plus 70 mmol KHCO3 (HP+K), and HP plus 70 mmol NaCl (HP+Na). Protein was casein:lactalbumin (80:20), except for the treatments with added sodium chloride, to which only casein was added. In study 2, the effects of HP and HP plus 50 mmol KHCO3 (HP+K) were compared with those of MP or MP plus 7.5 mmol phosphate (MP+Pi), equaling the additional phosphate of HP, in 10 adult men. Subjects completed all treatments in random order. In study 1, the peak of calcium excretion was at 3 h for all treatments, except for HP+K, which indicated an acute hypocalciuric effect of potassium. Unexpectedly, there was no hypercalciuric effect of adding sodium chloride, nor was urine sodium increased. In study 2, calcium excretion was significantly higher with HP than with MP+Pi but not with MP at 3 h, indicating an acute hypercalciuric effect of protein alone. A hypocalciuric effect of potassium (HP+K compared with HP) but not of phosphate (MP compared with MP+Pi) was seen. An acute load test measuring changes 3 h postload was appropriate for examining the calciuric effects of protein and potassium bicarbonate, but not those of sodium chloride or phosphate in adults.     

Regions of the haptoglobin 5' flanking gene sequence show different binding affinities to nuclear matrix proteins during the acute phase response

The effect of the acute phase response on the affinity of binding between nuclear matrix proteins and the rat haptoglobin (Hp) gene region was examined. Nuclear matrices isolated from acute phase livers were enriched with the 5' Hp gene flanking region (-705/+159), but not with the spliced, protein-coding sequence. Reassociation experiments with isolated nuclear protein matrix spheres and end-labeled fragments I (-146/+156), II (-146/-541), and III (-541/-705) revealed that the matrix proteins displayed an increased binding potential during the acute phase response for all of the examined regions, this being most pronounced for fragment II. BAL 31 digestion of fragment II showed that the sequence element that was responsible for the increased association with nuclear matrix proteins during the acute phase response was a tract of 38 adenine bases. The DNA region established stable associations with nuclear lamin B (67 kDa, pI 5.7) in the controls, and with lamins A (69 kDa, pI 7.0), B, isoforms of lamin C (62 kDa, pI 6.55-6.95), and a 55-kDa (pI 5.9) polypeptide during the acute phase response. Sequence ABC (-165/-56), which overlaps fragments I and II and represents the Hp cis-acting element, did not bind to the non-histone nuclear matrix proteins.     

Anchoring of c-myc on nuclear matrix proteins in process of mouse thymic T lymphocyte proliferation induced by ConA

Epidemiologic investigations of the Latin America cholera epidemic have repeatedly implicated untreated drinking water and water touched by hands during storage as important vehicles for disease transmission. To prevent such transmission, we provided a new narrow-mouthed, plastic, water storage vessel and 5% calcium hypochlorite solution for home disinfection of stored water to a Bolivian Aymara Indian community at risk for cholera. We evaluated acceptance of this intervention and its effect on water quality. Each of 42 families in the study obtained water from a household well; fecal coliform bacteria were found in water from 39 (93%) of 42 wells and 33 (79%) of 42 usual water storage vessels. One group of families received the special vessels and chlorine (group A), a second received only the special vessels (group B), and a third served as a control group (group C). Water samples collected every three weeks from group A special vessels had lower geometric mean fecal coliform colony counts (P < 0.0001) and lower geometric mean Escherichia coli colony counts (P < 0.0001) than water from group B or C vessels. Adequate levels of free chlorine persisted in these vessels for at least 5 hr. The special vessels and chlorine solution were well accepted and continued to be used for at least six months. Use of the vessel and chlorine solution produced drinking water from nonpotable sources that met World Health Organization standards for microbiologic quality.     

Changes in skeletal metabolism in pubertal girls can be revealed by biochemical parameters of calcium metabolism and bone turnover

Biochemical changes related to skeletal turnover in puberty were investigated in a sample of 67 girls aged 8-14 years. The following biochemical parameters were measured in serum: total calcium, phosphate, magnesium, total alkaline phosphatase, osteocalcin, and calcium and hydroxyproline in the second morning urine. Thirty-five premenarchal girls (8-11 years) had significantly lower serum calcium, and higher alkaline phosphatase and phosphate than those menstruating regularly (N = 32, 12-14 years). A statistically significant negative correlation of serum parameters and age was found for phosphate and alkaline phosphatase in all subjects, and for calcium and magnesium only in the premenarchal girls. These results indicated the more intensive processes of skeletal metabolism occurring in prepubertal age and early puberty to reflect in basic biochemical parameters of calcium and bone metabolism. Analysis of correlation between biochemical parameters showed alkaline phosphatase and phosphate to correlate positively with hydroxyproline excretion and negatively with urinary calcium in all subjects. In the subjects after menarche, osteocalcin correlated with alkaline phosphatase and phosphate. Thus, biochemical parameters indirectly reflected physiologic changes occurring with bone turnover in puberty. Variations in bone turnover during puberty, including a more pronounced bone formation during prepubertal or early stages, can be indirectly observed through biochemical parameters related to calcium and bone metabolism. Investigations of skeletal growth and puberty would benefit from specific markers of bone remodeling and "basic" biochemical parameters, as it might disclose subtle metabolic relationships.     

Inhibition of fear-potentiated startle can be detected after the offset of a feature trained in a serial feature-negative discrimination.

Using the fear-potentiated startle paradigm in rats, 4 experiments examined whether the inhibitory effect of a feature is evident after its offset following serial feature-negative discrimination training (A+ and X?→?A–). When startle probes were presented shortly after the offset of X on X?→?A test trials, the inhibitory properties of X were observed immediately after its offset. Furthermore, trace reinforcement of X (X?→?+), but not delay reinforcement (X+), disrupted the ability of X to inhibit fear-potentiated startle on X?→?A trials. Trace conditioning to X was also retarded after A+ and X?→?A– training. These results suggest that the inhibitory properties of the serially trained feature are present after its offset and raise the possibility that either temporal information regarding nonreinforcement or poststimulus attributes of X acquire inhibitory properties. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Changes in the distribution of short-term exposure concentration with different averaging times

PURPOSE: To express for educational purposes, different and often opposing views on controversial topics of current interest in vascular radiology. And to explore whether or not consensus can be reached on any of these controversial issues. METHOD: Panel discussion among experts in the field. With the help of a moderator, controversial topics were introduced for discussion. For each topic the moderator underscored the principal issue and asked relevant questions. All panelists were asked to express their views on all issues. The following issues were discussed: Atherectomy vs balloon angioplasty; Balloon angioplasty vs stents; Digital vs analog Imaging; TIPS as an emergency procedure; Regional thrombolysis; MRA vs conventional angiography; Laser angioplasty; Carbon dioxide angiography; Lymphangiography vs CT; The training of surgeons in angiography. The panelists were urged to avoid extensive references to the literature but rather to express their own personal opinions based on experience and practice. For each topic an attempt was made to arrive at consensus. RESULTS: Ten issues were presented for debate and discussion. Despite divergent opinions it was possible and relatively painless to reach consensus on seven controversies. On the remaining issues the panelists agreed to disagree. However even when there was disagreement certain trends became apparent. Local considerations and resource availability accounted for varied approaches to the solution of certain controversies. CONCLUSIONS: A panel discussion among experts may be a useful way to address controversies for educational purposes. Despite divergence of opinions consensus may be reached, or in absence of consensus general trends may become apparent.     

Expression of two different antiphagocytic M proteins by Streptococcus pyogenes of the OF+ lineage

All clinical isolates of Streptococcus pyogenes (group A streptococcus) share the ability to resist phagocytosis and grow in human blood. In many strains, this property is due to the expression of a single antiphagocytic M protein, while other strains express more than one M-like molecule, of which the role in phagocytosis resistance is unclear. In particular, all S. pyogenes strains of the OF+ lineage, representing approximately half of all isolates, express two M-like proteins, Mrp and Emm, which are immunologically unrelated. These two proteins bind different ligands that have been implicated in phagocytosis resistance: Mrp binds fibrinogen and Emm binds the complement inhibitor C4BP. Using a clinical isolate of the common serotype 22, we created mutants affected in the mrp and emm genes and characterized them in phagocytosis experiments and by electron microscopy. A double mutant mrp-emm- showed strongly decreased resistance to phagocytosis, while mrp- and emm- single mutants grew well in blood. However, optimal growth required the expression of both Mrp and Emm. Experiments in which coagulation was inhibited using the specific thrombin inhibitor, hirudin, rather than heparin, indicated that Emm is more important than Mrp for resistance to phagocytosis. Tuftlike surface structures typical for S. pyogenes were still present in the mrp-emm- double mutant, but not in a mutant affected in the regulatory gene mga, indicating that the presence of these surface structures is not directly correlated to phagocytosis resistance. Our data imply that OF+ strains of S. pyogenes express two antiphagocytic M proteins with different ligand-binding properties.     

Translation of RNAIII, the Staphylococcus aureus agr regulatory RNA molecule, can be activated by a 3''-end deletion

A case of enteropathy associated T-cell lymphoma (EATCL) in a 62-year-old female with a previous history of coeliac disease, complicated during the clinical course by massive blood and tissue eosinophilia is described. The patient's serum contained a factor capable of stimulating the in vitro growth of eosinophilic colonies (CFU-Eo), that was absent in the serum of normal donors. We suggest that such factor was Interleukin-5 (IL-5), as indicated by the presence in the monoclonal tumor T cells of IL-5 encoding mRNA, usually absent in the normal enterocytes of the jejunum.     

beta-subunit of nuclear pore-targeting complex (importin-beta) can be exported from the nucleus in a Ran-independent manner

The nuclear export of importin-alpha is mediated by CAS, which is related to importin-beta, whereas the mechanism for the export of importin-beta remains unclear. In this study, we demonstrate that the nuclear export of importin-beta is mediated by the nuclear pore complex-binding domain of this molecule. Insensitivity to leptomycin B indicates that its export is not mediated by a leucine-rich nuclear export signal-specific receptor, CRM1. Furthermore, the nuclear export of importin-beta was not inhibited by co-injection with a GTPase-deficient Ran mutant (G19V). The cell line tsBN2 contains a temperature-sensitive point mutation in the RCC1 gene, which encodes a guanine nucleotide exchange factor of Ran. At the nonpermissive temperature, importin-beta was exported from the nucleus of these cells, even when RanGAP1, a GTPase-activating protein for Ran, was co-injected. These results not only provide support for the view that Ran-dependent GTP hydrolysis is not required for the nuclear export of importin-beta but also indicate that nuclear RanGTP is not essential for its export. As a result, we propose that importin-beta can be recycled from the nucleus alone in a Ran-independent manner.     

A comparison of the mechanical properties and microstructures of intermetallic matrix composites fabricated by two different methods

The tensile properties of SCS-6 SiC fiber-reinforced Ti-24Al-11Nb (at. pct) have been measured over the past several years by a number of investigators. These composites have been fabricated by different techniques and tend to exhibit a large amount of scatter in the longitudinal tensile properties. To date, it is not known if one optimized fabrication method provides composites with improved mechanical properties over those produced by other optimized methods, since carefully controlled experiments have not been performed to determine this. Thus, the purpose of the present study was to compare the longitudinal tensile strengths of SCS-6 SiC/ Ti-24Al-11Nb composites that had been fabricated by the powder-cloth method and the lowpressure plasma spray (LPPS) method. In this study, the same lots of matrix powder and reinforcing fiber were used for fabricating the composites. It was determined that the powder-cloth and plasma spray methods produced composites having very similar tensile properties. Both fabrication methods induced damage in a small percentage of fibers, which manifested itself in the form of bimodal Weibull distributions of extracted fiber strengths. It appeared that the particular lot of SCS-6 fiber used in fabricating both types of composites was more susceptible to fabrication damage than those used in previous studies. This article also shows the dramatic effect that different handling and testing techniques can have on measured fiber strengths.     

Convulsions due to increased permeability of the blood-brain barrier in experimental cerebral malaria can be prevented by splenectomy or anti-T cell treatment

Experimental cerebral malaria (ECM) can be induced in C57B1 mice by infection with Plasmodium berghei K173 parasites. Behavioral changes shortly before they die of ECM may reflect disturbance of the integrity of the blood-brain barrier (BBB). Folic acid elicits strong convulsive activity if the permeability of the BBB is increased. Administration of folic acid to mice during development of ECM induced convulsions. Interventions known to prevent fatal outcome from ECM, such as splenectomy or treatment with anti-CD4 or anti-CD8 monoclonal antibodies, also prevented sensitivity to folic acid-induced convulsions. In addition, infected mice with ECM and sensitive to folic acid-induced convulsions, recovered from this sensitivity after treatment with anti-T cell antibodies within 4 h. These data suggest that disturbance of the permeability of the BBB can be reversed and depends on the involvement of T cells.     

Intronic U14 snoRNAs of Xenopus laevis are located in two different parent genes and can be processed from their introns during early oogenesis

U14 is a member of the rapidly growing family of intronic small nucleolar RNAs (snoRNAs) that are involved in pre-rRNA processing and ribosome biogenesis. These snoRNA species are encoded within introns of eukaryotic protein coding genes and are synthesized via an intron processing pathway. Characterization of Xenopus laevis U14 snoRNA genes has revealed that in addition to the anticipated location of U14 within introns of the amphibian hsc70 gene (introns 4, 5 and 7), additional intronic U14 snoRNAs are also found in the ribosomal protein S13 gene (introns 3 and 4). U14 is thus far a unique intronic snoRNA in that it is encoded within two different parent genes of a single organism. Northern blot analysis revealed that U14 snoRNAs accumulate during early oocyte development and are rapidly expressed after the mid-blastula transition of developing embryos. Microinjection of hsc70 pre-mRNAs into developing oocytes demonstrated that oocytes as early as stages II and III are capable of processing U14 snoRNA from the pre-mRNA precursor. The ability of immature oocytes to process intronic snoRNAs is consistent with the observed accumulation of U14 during oocyte maturation and the developmentally regulated synthesis of rRNA during oogenesis.     

Expression of glyoxylate cycle genes in cucumber roots responds to sugar supply and can be activated by shading or defoliation of the shoot

Nuclear localization signals (NLSs) are short peptides required for nuclear transport of karyophilic proteins. We review in this paper how the nuclear targeting property of NLS peptides has been taken advantage of to enhance the efficiency of nuclear uptake of transgene DNA in zebrafish and how it may improve the efficiency of transgenesis in this species. Synthetic NLS peptides can bind to plasmid DNA by ionic interactions. Cytoplasmic injection of DNA-NLS complexes in zebrafish eggs enhances the rate and the amount of plasmid DNA taken up by embryonic nuclei. Nuclear import of DNA-NLS complexes has been duplicated in vitro and exhibits energetic and cytosolic requirements similar to those for nuclear protein import. Furthermore, binding NLSs to DNA increases expression frequency of the transgene. We suggest that NLS peptides may constitute a valuable tool to improve the efficiency of transgenesis in zebrafish and other species.     

Resistance to paclitaxel mediated by P-glycoprotein can be modulated by changes in the schedule of administration

OBJECTIVE: The objective of this study was to examine the occurrence of ovarian endometriosis in epithelial ovarian cancer in Japan. METHOD: The presence of ovarian endometriosis was determined by reviewing the sections of resected specimens in 172 epithelial ovarian cancers. RESULTS: The incidence of ovarian endometriosis in ovarian cancer (14.5%) was higher than that in Western countries. The rank order of incidence of endometriosis in each histologic type was clear cell (40.6%)>endometrioid (23.1%)>serous (8.7%)>mucinous (2.9%). The incidence in serous type was higher when compared with that reported in Western countries. The higher incidence of endometriosis in Japan can be explained by a greater proportion of clear cell type, comprising 18.6% of all the cases and a higher incidence of endometriosis in the serous type. CONCLUSION: The association of ovarian endometriosis with epithelial ovarian cancer was more frequently found in Japan.     

Analysis of the low molecular weight nuclear polypeptide by isoelectrofocusing of nuclear proteins first separated by gel electrophoresis in SDS

Some of the nuclear proteins, especially those tightly bound with nucleic acids, show a tendency to aggregate and some of them are also hydrophobic. Thus, SDS-containing buffers are useful for their analysis. The method of isoelectrofocusing of proteins separated by polyacrylamide gel electrophoresis in the presence of SDS was used for the analysis of the nuclear proteins. First, SDS-polyacrylamide gel electrophoresis of total chromatin proteins was conducted. The sample of the examined nuclear proteins after SDS-gel electrophoresis was isolated as a gel slice (thus corresponding to the particular molecular weight range) and used for isoelectrofocusing. This approach has been used for the analysis of about 19 kD nuclear polypeptide, tightly bound with DNA. The isoelectric point of this polypeptide has been estimated as 5.3.     

"BEP" RNAs and proteins are situated in the animal side of sea urchin unfertilized egg, which can be recognized by female pronuclear localization

Microsurgery experiments demonstrate that the animal side of the unfertilized sea urchin Paracentrotus lividus egg coincides with the side of the egg pronucleus location. It is demonstrated by means of in situ hybridization and immunostaining of whole mounts of animal or vegetal halves that the previously identified bep 1 and bep4 RNAs and their proteins are located in the animal part of the unfertilized egg and much less in the vegetal part. The addition of Fabs against BEP1 and BEP4 causes exogastrulation.