Scrapie associated fibrils in found dead sheep.
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Animals↗
Biomedical subjects
A M Clark
Publications and source records attributed to A M Clark.
LagC is required for cell-cell interactions that are essential for cell-type differentiation in Dictyostelium.
Strain AK127 is a developmental mutant of Dictyostelium discoideum that was isolated by restriction enzyme-mediated integration (REMI). Mutant cells aggregate normally but are unable to proceed past the loose aggregate stage. The cloned gene, lagC (loose aggregate C), encodes a novel protein of 98 kD that contains an amino-terminal signal sequence and a putative carboxy-terminal transmembrane domain. The mutant strain AK127 shows no detectable lagC transcript upon Northern analysis, indicating that the observed phenotype is that of a null allele. Expression of the lagC cDNA in AK127 cells complements the arrest at the loose aggregate stage, indicating that the mutant phenotype results from disruption of the lagC gene. In wild-type cells, lagC mRNA is induced at the loose aggregate stage and is expressed through the remainder of development. lagC- null cells aggregate but then disaggregate and reaggregate to form small granular mounds. Mature spores are produced at an extremely low efficiency (< 0.1% of wild type), appearing only after approximately 72 hr, whereas wild-type strains produce mature spores by 26 hr. lagC- null cells accumulate reduced levels of transcripts for the prestalk-enriched genes rasD and CP2 and do not express the DIF-induced prestalk-specific gene ecmA or the cAMP-induced prespore-specific gene SP60 to significant levels. In chimeric organisms resulting from the coaggregation of lagC- null and wild-type cells, cell-type-specific gene expression is rescued in the lagC- null cells; however, lagC- prespore cells are localized to the posterior of the prespore region and do not form mature spores, suggesting that LagC protein has both no cell-autonomous and cell-autonomous functions. Overexpression of lagC from an actin promoter in both wild-type and lagC- cells causes a delay at the tight aggregate stage, the first stage requiring LagC activity. These results suggest that the LagC protein functions as a nondiffusible cell-cell signaling molecule that is required for multicellular development.
Actins↗
Leptospira hardjo infection in sheep.
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Abortion, Veterinary↗
Antimicrobial properties of alkaloids from Xanthorhiza simplicissima.
The organic extract of the whole plant Xanthorhiza simplicissima was found to exhibit good activity against the AIDS-related opportunistic pathogens Candida albicans, Cryptococcus neoformans, and Mycobacterium intracellularae. Bioassay-directed fractionation of the extract led to the isolation of the known alkaloid berberine as the major active component. A second alkaloid of the isohomoprotoberberine family, puntarenine, was isolated from this plant family for the first time. Puntarenine also showed marginal activity against the dermatophytic fungus Trichophyton mentagrophytes and the yeast Saccharomyces cerevisiae.
Alkaloids↗
Spatial and temporal expression of the Dictyostelium discoideum G alpha protein subunit G alpha 2: expression of a dominant negative protein inhibits proper prestalk to stalk differentiation.
Previous results have shown that the G alpha protein subunit G alpha 2 is required for aggregation in Dictyostelium discoideum and is essential for coupling cell-surface cAMP receptors to downstream effectors in vivo during this stage of development. G alpha 2 expresses at least four distinct transcripts that are differentially regulated during development; two of the transcripts are expressed exclusively in the multicellular stages and their expression is restricted to prestalk cells. We partially dissected the G alpha 2 promoter and identified a component that is expressed exclusively during the multicellular stages using luciferase gene fusions. When this promoter region is coupled to lacZ, beta-gal expression is restricted to the multicellular stages and localized in prestalk cells with a pattern similar to that of the ecmA prestalk-specific promoter. We show that expression in wild-type cells of the G alpha 2 mutant protein [G alpha 2(G206T)] during the early stages of development blocks aggregation and cAMP-mediated activation of adenylyl cyclase and guanylyl cyclase, suggesting it functions as a dominant negatively active G alpha subunit. When this mutant G alpha protein is expressed from the ecmA prestalk-specific promoter, abnormal stalk differentiation during culmination is observed. Expression of the mutant G alpha 2 from the SP60 prespore promoter or wild-type G alpha 2 from either the ecmA or the SP60 promoter results in no detectable phenotype. The results suggest that G alpha 2 plays an essential role during the culmination stage in prestalk cells and may mediate cAMP receptor activation of these processes during multicellular development.
Amino Acid Sequence↗
A primate model for chancroid.
Adult pigtailed macaques (Macaca nemestrina) were evaluated for their usefulness as a primate model for chancroid. To initiate infection, 10(7)-10(8) cfu of Haemophilus ducreyi were inoculated into the foreskins of 5 adult males and into the vaginal labia of 4 adult females. Lesions developed in the male macaques that were similar in appearance, histopathologic changes, and progression to those of human disease, including the development of ulcers 6-12 days after infection. In addition, H. ducreyi could be recovered from the lesions up to 20 days after inoculation, humoral antibodies were induced beginning 1 week after inoculation, and inguinal lymphadenopathy was noted in 4 of the 5 males. None of the 4 female macaques inoculated with the same preparation of live H. ducreyi developed comparable lesions. Thus, experimental chancroid in adult male macaques closely resembles human disease and should be useful for future studies of the pathogenesis of chancroid.
Animals↗
The obstetric outcome of singleton pregnancies following in-vitro fertilization/gamete intra-fallopian transfer.
The present study compares 465 singleton live deliveries from in-vitro fertilization/gamete intra-Fallopian transfer (IVF/GIFT) pregnancies with a large control population to evaluate the incidence of pre-term delivery and small for gestational age (SGA) or very small for gestation age (VSGA) babies resulting from IVF/GIFT pregnancies. Overall the incidence of SGA or VSGA from an IVF/GIFT pregnancy is higher than from the normal obstetric population (SGA odds ratio 1.76, 95% confidence interval (CI): 1.38-2.25 and VSGA odds ratio 1.61, 95% CI: 1.05-2.46) particularly among primiparous women (SGA odds ratio 1.99, 95% CI: 1.25-3.16 and VSGA odds ratio 1.97, 95% CI: 1.49-2.62). After stratifying by the cause of infertility, only women with unexplained infertility had a significantly higher proportion of SGA/VSGA babies. There was a significantly higher incidence of pre-term deliveries among the young primiparae (odds ratio 5.02, 95% CI: 3.09-8.13). Thus the excess risk of delivering a SGA/VSGA baby and pre-term delivery from an IVF/GIFT pregnancy seems to be largely confined to women with unexplained infertility and young primiparae.
Adult↗
Embryo viability and implantation.
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Embryo Implantation↗
Changes in the messenger ribonucleic acid for insulin-like growth factor-I and -II in the porcine testis during and between two waves of testicular development.
The boar testis was used as a model for examining the possible role of production of the insulin-like growth factors (IGF) in steroidogenesis and/or testicular growth as testicular development occurs in waves. Blood and testes were sampled from boars at different ages (100-102 days of gestation; 7, 19, and 30 days; and 10 and 25 wk), selected to occur during and between the last two waves of testicular development. Serum was analyzed for testosterone and RNA was extracted from the testes for Northern and dot-blot analysis of IGF mRNA. Testosterone concentrations declined (p = 0.01) from 7 days to 10 wk of age and rebounded at 25 wk, indicating completion of the second and third waves of testicular development. The quantity of testicular mRNA for IGF-I increased gradually with age, and that for IGF-II decreased. We therefore conclude that regulation of the expression of the mRNAs for IGF-I and -II in the pig testis is not a function of either the waves of testicular development or the level of steroidogenesis.
Actins↗
Regulatory role of the G alpha 1 subunit in controlling cellular morphogenesis in Dictyostelium.
To determine the function of the Dictyostelium G alpha 1 subunit during aggregation and multicellular development, we analyzed the phenotypes of g alpha 1 null cells and strains overexpressing either wild-type G alpha 1 or two putative constitutively active mutations of G alpha 1. Strains overexpressing the wild-type or mutant G alpha 1 proteins showed very abnormal culmination with an aberrant stalk differentiation. The similarity of the phenotypes between G alpha 1 overexpression and expression of a putative constitutively active G alpha 1 subunit suggests that these phenotypes are due to increased G alpha 1 activity rather than resulting from a non-specific interference of other pathways. In contrast, g alpha 1 null strains showed normal morphogenesis except that the stalks were thinner and longer than those of wild-type culminants. Analysis of cell-type-specific gene expression using lacZ reporter constructs indicated that strains overexpressing G alpha 1 show a loss of ecmB expression in the central core of anterior prestalk AB cells. However, expression of ecmB in anterior-like cells and the expression of prestalk A-specific gene ecmA and the prespore-specific gene SP60/cotC appeared normal. Using a G alpha 1/lacZ reporter construct, we show that G alpha 1 expression is cell-type-specific during the multicellular stages, with a pattern of expression similar to ecmB, being preferentially expressed in the anterior prestalk AB cells and anterior-like cells. The developmental and molecular phenotypes of G alpha 1 overexpression and the cell-type-specific expression of G alpha 1 suggest that G alpha 1-mediated signaling pathways play an essential role in regulating multicellular development by controlling prestalk morphogenesis, possibly by acting as a negative regulator of prestalk AB cell differentiation. During the aggregation phase of development, g alpha 1 null cells display a delayed peak in cAMP-stimulated accumulation of cGMP compared to wild-type cells, while G alpha 1 overexpressors and dominant activating mutants show parallel kinetics of activation but decreased levels of cGMP accumulation compared to that seen in wild-type cells. These data suggest that G alpha 1 plays a role in the regulation of the activation and/or adaptation of the guanylyl cyclase pathway. In contrast, the activation of adenylyl cyclase, another pathway activated by cAMP stimulation, was unaffected in g alpha 1 null cells and cell lines overexpressing wild-type G alpha 1 or the G alpha 1 (Q206L) putative dominant activating mutation.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
Induction of two alternatively spliced evi-1 proto-oncogene transcripts by cAMP in kidney cells.
The evi-1 proto-oncogene is normally predominantly expressed in the kidney. We report here that evi-1 transcripts are also abundant in foetal kidney and expression is retained in primary kidney cell cultures. However available kidney cell lines express low or no evi-1 mRNA. In the human renal cell carcinoma cell line, A704, evi-1 is inducible approximately 16-fold by elevating intra-cellular cAMP levels with either forskolin or dibutyryl cAMP. TPA down-regulates evi-1 mRNA production and blocks forskolin mediated induction. Similar effects are seen in NIH3T3 cells and primary kidney cell cultures. Induction of evi-1 gene expression by forskolin does not alter the ratio of full length and an alternatively spliced transcript which encodes a protein lacking two repeats of the zinc finger motif. Potential regulation of evi-1 expression in kidney by hormones which modulate intra-cellular cAMP levels suggest that it can respond to environmental cues which might be important to the normal physiological role of this protein in kidney differentiation, development and function.
3T3 Cells↗
Micronuclei in adult and foetal mice exposed in vivo to heliotrine, urethane, monocrotaline and benzidine.
The level of clastogenic damage in mice following in vivo adult and transplacental exposure to benzidine, heliotrine, monocrotaline and urethane was compared using the micronucleus (MN) assays of polychromatic erythrocytes (PCE) found in mouse adult bone marrow and foetal liver. Saline was used as a negative control. Swiss-albino mice (gestation 17-19 days) were given a single acute intraperitoneal injection. Animals sacrificed 21 h after 0.75 LD50 of each chemical had significantly increased frequencies of micronucleated PCE in adult and foetal tissues (p < 0.05). Heliotrine resulted in the largest increases in mean value of micronucleated PCE, of 5.4% in adult bone marrow and 6.9% in foetal liver compared to control values of 0.7% and 0.9% for adult and foetal tissue respectively (p < 0.005). The induction of MN was significantly higher in foetal than adult cells for 0.75 LD50 monocrotaline and benzidine after 21 h (p < 0.0005), and for one LD50 heliotrine at 12, 24 and 30 h after injection in a time-response study (p < 0.02). Induction of MN following LD50 heliotrine showed peak MN expression in PCE at 18 h after injection for adult bone marrow (2.6%) and at 24 h for foetal liver (7.9%). The majority of micronucleated PCE in adult and foetal tissue contained only one MN (mean 74% in mice treated with chemicals and 91% in untreated mice). This study demonstrated the sensitivity of the transplacental MN assay for detecting clastogenic damage in mice induced by chemicals and their metabolites. The importance of monitoring indicators of toxicity was highlighted by changes in erythrocyte population composition and the presence of basophilic stippling following exposure to LD50 heliotrine.
Animals↗
Determinants of non-spatial working memory deficits in rats given intraventricular infusions of the NMDA antagonist AP5.
Two series of experiments using rats assessed the effects of intraventricular administration of the NMDA antagonist AP5 on performance of non-spatial working memory tasks. The first series used a continuous delayed non-matching to sample (DNMS) design; the second series used a discrete trial delayed matching to sample (DMS) design. Performance was assessed at retention intervals ranging from approximately 5 to 90 sec. The subjects had acquired the behavioural tasks before drug testing commenced. In the DNMS series, minipumps containing vehicle, 5, 10 or 15 nM D-AP5 were implanted. Every 10 days, each rat's minipump was removed and replaced with a fresh pump containing a new drug dose in a counterbalanced design, so that all rats were tested under all four conditions. There were no drug effects on performance at any retention interval. In the DMS series, there were three different basic task variants. Minipumps filled either with 15 mM D-AP5 or vehicle solution were implanted. Vehicle rats performed at approximately pre-operative levels; AP5 rats were impaired only on task variants using repeated stimulus presentations within session. There was no interaction between retention interval and drug treatment. This pattern of results closely resembles that seen following hippocampectomy or fornicotomy, as would be expected if this drug, administered intraventricularly, selectively affected hippocampal function.
2-Amino-5-phosphonovalerate↗
Antimicrobial compounds from Petalostemum purpureum.
EtOH extracts of Petalostemum purpureum demonstrated antimicrobial activity against bacteria and fungi. Bioassay-directed fractionation led to the isolation of petalostemumol [1] as the active constituent. Its structure was determined by a single crystal X-ray diffraction analysis. A minor component designated petalostemumol G [3] is believed to be an artifact of the isolation procedure. Its structure was confirmed by a single crystal X-ray analysis of its pentamethylether 2. A number of derivatives of 1 and 3, including the Me- and benzylethers and acetates, were prepared.
Acetylation↗
Chemical composition and antimicrobial activity of essential oils of Jasonia candicans and J. montana.
The essential oils of the aerial parts of Jasonia candicans and J. montana were analyzed by gas chromatography-mass spectrometry (GC/MS) technique. Of twenty-one components identified in the volatile oil of J. candicans, intermediol was the main constituent. Fifty-eight components were characterized in the essential oil of J. montana. Camphor, borneol, bornyl acetate, chrysanthemol, intermediol, and 1,8-cineole were the major constituents in this oil. The two oils showed antibacterial activity against Bacillus subtilis. They also showed a marked antifungal activity against Trichophyton mentagrophytes, Cryptococcus neoformans, and Candida albicans.
Anti-Bacterial Agents↗
Gamete intra-fallopian transfer: outcome following the elective or non-elective replacement of two, three or four oocytes.
The outcome of 807 gamete intra-Fallopian transfer (GIFT) cycles following the elective or non-elective transfer of two, three or four oocytes has been retrospectively studied. Electively replacing either three or two oocytes did not reduce the clinical or ongoing pregnancy rate when compared with replacing four oocytes. The incidence of high-order multiple gestation (triplet or more) was significantly reduced by replacing fewer oocytes, but the occurrence of twin pregnancy was not altered. Lower pregnancy rates were found when the number of oocytes available for replacement was limited and non-elective replacement was performed. It is suggested, therefore, that a higher number of oocytes available may allow selection of higher quality oocytes for transfer. We conclude that the overall expectation of pregnancy from the GIFT procedure is high (30-40%) and the number of oocytes replaced should be two in order to minimize the risk of high-order multiple pregnancies. It is not clear whether increasing the number of oocytes transferred will benefit subjects who failed to become pregnant previously with GIFT, but limited data suggest that transferring large numbers of oocytes to women > 40 years does not improve the expectation of pregnancy.
Age Factors↗
Epidermis-specific transcripts. Nucleotide sequence of a full-length cDNA of EPI12, encoding a putative lipid transfer protein.
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Base Sequence↗
Diagnosis of Chlamydia trachomatis urethritis in men by polymerase chain reaction assay of first-catch urine.
To determine the accuracy of a recently developed polymerase chain reaction (PCR) urine assay to detect Chlamydia trachomatis urethral infection in men, we obtained urethral swabs and first-catch urine from 365 men attending a sexually transmitted diseases clinic. Thirty-three (9%) of the 365 men were infected with C. trachomatis as defined by urethral culture. Thirty-two of the 33 men with culture-positive urethral swabs also had PCR-positive urine assays. Of 332 patients with culture-negative urethral swabs, 325 had PCR-negative urine. Compared with chlamydia culture of urethral specimens, PCR assay of urine samples thus had a sensitivity of 97% and a specificity of 98%. The positive predictive value of the urine PCR assay was 82%, and the negative predictive value was 99%. Analysis of discrepant results indicated that six of seven PCR-positive, urethral culture-negative patients probably had chlamydial urethritis. All six patients had symptoms of urethritis and had either a positive urethral swab PCR or a positive urine PCR with a different amplification target. After resolution of discrepant results, (defining true positives as the 33 culture-positive patients and the 6 PCR-positive, culture-negative patients just described), the sensitivity and specificity of culture were 85% (33 of 39) and 100% (326 of 326), respectively. The revised sensitivity and specificity of PCR were 97% (38 of 39) and 99.7% (325 of 326), respectively. We conclude that this urine PCR assay provides a highly sensitive, noninvasive alternative method for the detection of C. trachomatis urethral infection in high-risk men attending a sexually transmitted diseases clinic. This assay could greatly facilitate the testing of larger numbers of male patients for chlamydial infection and should be studied in other settings.
Bacterial Outer Membrane Proteins↗