A cAMP-activated chloride channel in the plasma membrane of cultured human gastric cells (HGT-1)

Hydrochloric acid (HCl) secretion by gastric parietal cells involves an apical Cl^– conductance, the properties of which have not been defined. In the present study, forskolin and histamine [agonists that increase intracellular cyclic adenosine monophosphate (cAMP)], and dibutyryl cAMP, activated channels in previously quiescent cell-attached membrane patches on cultured human gastric cells (HGT-1). In the cell-attached configuration (Cl^–149 mmol/ 1 in bath and pipette), channels exhibited outward rectification, voltage dependence, inward current (–0.7 pA) at zero holding potential and a reversal potential of +24 mV, consistent with the presence of a Cl^– conductive pathway. In excised inside-out patches, channels (i) exhibited degrees of outward rectification and voltage dependence that were comparable to those seen in cell-attached patches, (ii) demonstrated a –21 mV shift of their reversal potential when bath Cl^– was decreased from 149 mmol/l to 53 mmol/l (calculated Cl^–:cation permeability ratio 171), and (iii) were highly sensitive to the Cl^– channel blocker diphenylamine-2-carboxylic acid (DPC, 10^–3 mol/l). This cAMP-activated Cl^– channel bears many similarities to other Cl^– channels within intestinal epithalia, and may represent the apical Cl^– channel operating in HCl-secreting gastric parietal cells.     

Genetic analysis and attribution of microbial forensics evidence

Because of the availability of pathogenic microorganisms and the relatively low cost of preparing and disseminating bioweapons, there is a continuing threat of biocrime and bioterrorism. Thus, enhanced capabilities are needed that enable the full and robust forensic exploitation and interpretation of microbial evidence from acts of bioterrorism or biocrimes. To respond to the need, greater resources and efforts are being applied to the burgeoning field of microbial forensics. Microbial forensics focuses on the characterization, analysis and interpretation of evidence for attributional purposes from a bioterrorism act, biocrime, hoax or inadvertent agent release. To enhance attribution capabilities, a major component of microbial forensics is the analysis of nucleic acids to associate or eliminate putative samples. The degree that attribution can be addressed depends on the context of the case, the available knowledge of the genetics, phylogeny, and ecology of the target microorganism, and technologies applied. The types of genetic markers and features that can impact statistical inferences of microbial forensic evidence include: single nucleotide polymorphisms, repetitive sequences, insertions and deletions, mobile elements, pathogenicity islands, virulence and resistance genes, house keeping genes, structural genes, whole genome sequences, asexual and sexual reproduction, horizontal gene transfer, conjugation, transduction, lysogeny, gene conversion, recombination, gene duplication, rearrangements, and mutational hotspots. Nucleic acid based typing technologies include: PCR, real-time PCR, MLST, MLVA, whole genome sequencing, and microarrays.     

NK cells developing in vitro from fetal mouse progenitors express at least one member of the Ly49 family that is acquired in a time-dependent and stochastic manner independently of CD94 and NKG2

NK cells developing in vitro from fetal progenitors in the presence of IL-2 are phenotypically and functionally indistinguishable from mature adult NK cells, with the exception that they generally lack surface expression of any of the Ly49 molecules that have previously been examined. Using two recently developed anti-Ly49 mAb, we show here that most of these NK cells in fact express high levels of at least one previously uncharacterized member of the Ly49 family, most likely Ly49E. Detailed kinetic and clonal analysis revealed that these Ly49 molecules were acquired in a progressive and stochastic manner independently of CD94 and NKG2. CD94 and NKG2 were both expressed early in NK cell development, sometimes in the absence of NK1.1, with CD94 invariably being expressed at two different levels. IL-4 differentially inhibited the expression of CD94 and Ly49 receptors, but had little or no effect on the expression of NKRP1 molecules.     

Functional stability of dorsolateral prefrontal neurons

Stable multiday recordings from the dorsolateral prefrontal cortex of 2 monkeys performing 2 Go/NoGo visual-discrimination tasks (one requiring well-learned responses, the other requiring learning) demonstrate that the majority of prefrontal neurons were "functionally stable". Recordings were made using a series of removable microdrives, each implanted for 3-6 mo, housing independently mobile electrodes. Action potential waveforms of 94 neurons were stable over 2-9 days; 66/94 (70%) of these cells responded each day, 22/94 (23%) never responded significantly, and 6/94 (6%) responded one day but not the next. Of 66 responsive neurons, 55 were selective for either Go or NoGo trials, individual stimuli, or eye movements. This selectivity was functionally stable (i.e., maintained) for 46/55 neurons across all recording days. Functional stability was also noted in terms of response strength (baseline firing rates compared with poststimulation firing rates) and event-related response timing. Two neurons with consistent responses in familiar testing conditions responded flexibly when the monkeys learned to make correct responses to novel stimuli. We conclude that the majority of prefrontal neurons were functionally stable during the performance of well-learned tasks. Such stability may be a general property of prefrontal neurons, given that neurons with 4 different types of task selectivity were found to be functionally stable. Conceptually similar studies based on long-term recordings in other cortical regions reached similar conclusions, suggesting that neurons throughout the brain are functionally stable.     

Induced luteolysis in the primate: rapid loss of luteinizing hormone receptors

The molecular mechanisms involved in luteolysis are still unclear in the primate. This study aimed to investigate the effect of induced luteolysis on the ovarian luteinizing hormone (LH) receptor and the steroidogenic enzyme, 3beta-hydroxysteroid dehydrogenase (3beta-HSD) in the marmoset monkey. Luteolysis was induced in the mid-luteal phase either directly by systemic prostaglandin F2alpha (PGF2alpha), or indirectly by LH withdrawal using systemic gonadotrophin releasing hormone antagonist (GnRHant) treatment. The LH receptor was studied by isotopic mRNA in-situ hybridization and in-situ ligand binding and 3beta-HSD expression was studied using isotopic mRNA in-situ hybridization and immunohistochemistry. Induced luteolysis was associated with a reduction in the expression of LH receptor (P < 0.0001) and 3beta-HSD mRNA, closely followed by a reduction in the LH receptor (P < 0.05) and 3beta-HSD protein concentrations within 24 h. There were no differences in the findings whether luteolysis was induced with PGF2alpha or GnRHant. This study shows that disparate mechanisms to induce luteolysis in the primate result in an identical rapid loss of the LH receptor and 3beta-HSD. In conclusion, induced luteolysis leads to rapid loss of the steroidogenic pathway in luteal cells.      

Origin of hematopoietic progenitors during embryogenesis

It has been widely accepted that hematopoietic and endothelial cell lineages diverge from a common progenitor referred to as the hemangioblast. Recently, analyses of the potential of progenitor cells purified from mouse embryos as well as embryonic stem cells differentiating in vitro resolved intermediate stages between mesodermal cells and committed precursors for hematopoietic and endothelial cell lineages. There are two distinct hematopoietic cell lineages which have different origins, i.e., primitive hematopoietic lineage derived from mesoderm or hemangioblasts and definitive hematopoietic lineage derived from endothelial cells. The endothelium is suggested to provide a milieu in which the definitive hematopoietic lineage acquires multiple potentials.     

Use of modified resistogram to type Candida albicans isolated from cases of vaginitis and from faeces in the same geographical area.

Ninety strains of Candida albicans were isolated from vaginal swabs taken by general practitioners from cases of vaginal candidosis. One hundred and nine strains were also isolated from 300 specimens of faeces received from outpatients living in the same geographical area. These strains were typed using a modification of the resistotyping method. We were unable to find any significant differences between the two populations of strains. This suggests that no particular resistotype has an increased ability to cause vaginitis.     

Effect of lymphoid Leukosis virus on performance of layer hens and the identification of infected chickens by tests on Meconia

The effect of subclinical infection with lymphoid leukosis virus (LLV) on the productivity traits of layer hens was investigated. In hens that shed gs-antigen of LLV to albumen, onset of sexual maturity was delayed by a mean of 11 days and the number of eggs laid was reduced -by 68 per hen up to 75 weeks of age. Shedding hens laid on average 2 g lighter eggs and of lesser specific gravity. Thirty-four % less chickens were obtained in the reproduction programmes from LLV-shedders in comparison with non-shedders. LLV had no significant effect on fertility and hatchability. Reduced egg-related performance was only directly related to LLV-shedding and dams' shedding status had no effect on the egg-related performance of their LLV-free progeny. Dams' shedding status, however, correlated with higher mortality (10% higher) among their progeny. The percentage of non-layers was also higher in progeny of LLV-shedders. Meconia were highly suitable samples for identifying both transmitting dams and infected chickens but only if the test for infectious virus was performed. ELISA on meconia was less reliable than the test for virus and therefore is not recommended for the detection of residual of infected chickens in the flocks selected for reduced gs-antigen shedding.     

Androgen receptor YAC transgenic mice carrying CAG 45 alleles show trinucleotide repeat instability

X-linked spinal and bulbar muscular atrophy (SBMA) is caused by a CAG repeat expansion in the first exon of the androgen receptor (AR) gene. Disease-associated alleles (37-66 CAGs) change in length when transmitted from parents to offspring, with a significantly greater tendency to shift size when inherited paternally. As transgenic mice carrying human AR cDNAs with 45 and 66 CAG repeats do not display repeat instability, we attempted to model trinucleotide repeat instability by generating transgenic mice with yeast artificial chromosomes (YACs) carrying AR CAG repeat expansions in their genomic context. Studies of independent lines of AR YAC transgenic mice with CAG 45 alleles reveal intergenerational instability at an overall rate of approximately 10%. We also find that the 45 CAG repeat tracts are significantly more unstable with maternal transmission and as the transmitting mother ages. Of all the CAG/CTG repeat transgenic mice produced to date the AR YAC CAG 45 mice are unstable with the smallest trinucleotide repeat mutations, suggesting that the length threshold for repeat instability in the mouse may be lowered by including the appropriate flanking human DNA sequences. By sequence-tagged site content analysis and long range mapping we determined that one unstable transgenic line has integrated an approximately 70 kb segment of the AR locus due to fragmentation of the AR YAC. Identification of the cis - acting elements that permit CAG tract instability and the trans -acting factors that modulate repeat instability in the AR YAC CAG 45 mice may provide insights into the molecular basis of trinucleotide repeat instability in humans.      

Concentration and turnover of intraperitoneal hyaluronan during inflammation

Aseptic peritonitis was induced in rabbits by intraperitoneal injection of irritating agents, mainly starch suspensions. The inflammatory response was followed in the peritoneal lavage fluid by cell counts (average increase about 800-fold the first day) and hyaluronan concentration (average increase about 200-fold on the second and third days). The turnover rate of hyaluronan was studied by injecting tritium-labeled hyaluronan intraperitoneally and by following the appearance of tritiated water in serum. In control animals given trace amounts of hyaluronan, half-lives of 1–14 h were recorded. When the labeled polysaccharide had been mixed with 10 mg/ml of unlabeled hyaluronan, the half-life was approximately one day. Rabbits with ongoing peritonitis exhibited half-lives between 1 and 16 h. It was concluded that there was a large individual variation in uptake kinetics, that the removal process could be receptor mediated, and that the increase in intraperitoneal hyaluronan in peritonitis mainly was due to an increased production of the polysaccharide rather than a decreased rate of removal.