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PubMed2004

Predictors of severity and necrosis in acute pancreatitis.

C-reactive protein remains the single standard biochemical marker for predicting the severity of AP. Because the combination of clinical-physiological scores and CRP provide good information at 48 hours, research has focused on the predictive ability of various markers when applied in the initial 24 hours after admission to the hospital. After detailed review of the literature, the authors conclude that there is no single tool that serves as the optimal predictor of severity. There are, however, data that support the use of certain tests to improve upon the clinician's early predictive ability on the subsequent course of AP. These include an APACHE II score greater than 7 and IL-6 at the time of admission, and urine TAP, urine trypsinogen-2, and serum PMN elastase at 24 hours (Table 4). These markers only will be able to help the clinician's predictive ability if they can be performed locally and if the results can be available ina timely manner. Future research should focus on promising markers such as procalcitonin, IL-8, IL-I ra, sTNFR, CAPAP, PLA-2, novel markers, and the combined use of more than one marker. The conventional research approach in predicting severity used in the last 15 years has limitations and appears to have reached its maximal potential. Novel conceptions and approaches, such as identification of genetic polymorphisms that predispose to severe course and complications of AP or other approaches are needed for a quantum step forward.

Acute Disease↗
PubMed2003

The effects of HHV-8 vMIP-II on SIVmac251 infection and replication competent and incompetent SIVmac239Delta3 vectors.

Human herpesvirus type 8 vMIP-II has one of the broadest ranges of chemokine receptor binding and therefore a multiplicity of biologic effects, both immunologic and antiviral. These properties make vMIP-II an attractive effector gene to be expressed from gene therapy vectors. The present studies were concerned with both therapeutic approaches: (1) an anti-simian immunodeficiency virus (SIV) biologic, and (2) an effector gene in SIV-based vectors. Regarding its antiviral properties, vMIP-II expressed from bacteria and SIV-based vectors bound the surface of CEMx174 cells and specifically suppressed SIVmac251 infection. A CCR3 monoclonal antibody partially inhibited vMIP-II binding, suggesting that both SIVmac251 and vMIP-II utilize a similar CCR3-like receptor for CEMx174 cell binding. Replication competent SIV-based vectors containing forward and reverse vMIP-II produced neither identifiable vMIP-II nor virions for the first 21 days. Virus replication occurred after this period. Significant sequence alterations in the forward vMIP-II containing replication competent vector transcripts were responsible for the failure of vMIP-II expression. The genetic basis for the initial failure to replicate virus and its later restoration was not determined but appeared in the II-PIMv containing vectors to coincide with deletions and compensatory rearrangements in nef 3' of the polypurine tract. Cells transfected with SIVmac239Delta3DeltaLTR-vMIP-II vectors expressed biologically active vMIP-II that bound CEMx174 cells and suppressed SIVmac251 infection. These data suggest that replication defective SIV vectors expressing immunobiolgic genes such as vMIP-II may prove useful in gene therapies, particularly in augmenting immune responses in chronically infected individuals.

Anti-HIV Agents↗
PubMed1995

Susceptibility of brook charr, Salvelinus fontinalis to the pathogenic haemoflagellate, Cryptobia salmositica, and the inheritance of innate resistance by progenies of resistant fish.

Eighty-seven of 175 laboratory raised F1 generation brook charr (Salvelinus fontinalis) were resistant to experimental Cryptobia salmositica infection. Susceptibility to the pathogen was not related to age nor to route of infection. Plasma of resistant charr lysed the parasite under in vitro conditions while those of susceptible fish did not. Sperm from 4 susceptible and 4 resistant male charr, from 8 F1 families, were used to fertilize eggs from 2 hatchery raised susceptible charr. Susceptibility of progenies (F2 families) was determined 4 weeks after inoculation of the parasite. All progenies (seven F2 families) of 4 susceptible males were susceptible to experimental infection and their plasma did not lyse the parasite. It was presumed that these susceptible progenies were homozygous recessive (rr), and that the resistant allele was dominant. All progenies (two F2 families) of 1 resistant male (presumed RR) and 2 susceptible females were all resistant to infection and their plasma lysed the parasite. The ratio of resistant to susceptible charr in 6 other F2 families from the other 3 resistant males (presumed Rr) and two susceptible females was about 1:1. The results suggest a single Mendelian locus which determines innate resistance to C. salmositica infection in brook charr and that it is possible to breed Cryptobia-resistant fish.

Alleles↗
PubMed2000

Rapid purification of transfected porcine muscle cells.

A detailed methodology is described for fluorescence-activated cell sorting (FACS) of porcine muscle cells that have been transfected to express green fluorescent protein (GFP). Cells are liberated from porcine skeletal muscle and primary cultures are transfected with DNA encoding GFP. Primary cultures are subjected to immunocytochemistry using a primary muscle-specific monoclonal antibody followed by incubation with a phycoerythrin-conjugated second antibody. Transfected myoblasts are sorted from fibroblasts using forward angle light scatter and ninety degree light scatter, phycoerythrin fluorescence, and GFP fluorescence. These procedures allow for isolation of genetically- engineered porcine muscle cells more rapidly than traditional clonal selection procedures. Consequently, FACS provides porcine myoblast populations that retain the majority of their replicative capacity and are not contaminated with non-myogenic cells.

Animals↗
PubMed2001

Y265H mutator mutant of DNA polymerase beta. Proper teometric alignment is critical for fidelity.

DNA polymerases have the unique ability to select a specific deoxynucleoside triphosphate from a pool of similarly structured substrates. One of these enzymes, DNA polymerase beta, offers a simple system to relate polymerase structure to the fidelity of DNA synthesis. In this study, a mutator DNA polymerase beta, Y265H, was identified using an in vivo genetic screen. Purified Y265H produced errors at a 40-fold higher frequency than the wild-type protein in a forward mutation assay. At 37 degrees C, transient kinetic analysis demonstrated that the alteration caused a 111-fold decrease in the maximum rate of polymerization and a 117-fold loss in fidelity for G misincorporation opposite template A. Our data suggest that the maximum rate of polymerization was reduced, because Y265H was dramatically impaired in its ability to perform nucleotidyl transfer in the presence of the correct nucleotide substrate. In contrast, at 20 degrees C, the mutant protein had a fidelity similar to wild-type enzyme. Both proteins at 20 degrees C demonstrate a rapid change in protein conformation, followed by a slow chemical step. These data suggest that proper geometric alignment of template, 3'-OH of the primer, magnesium ions, dNTP substrates, and the active site residues of DNA polymerase beta are important factors in polymerase fidelity and provide the first evidence that Tyr-265 is important for this alignment to occur properly in DNA polymerase beta.

Bacterial Proteins↗
PubMed1992

The emergence of simian/human immunodeficiency viruses.

Molecular evolutionary analyses strongly support the hypothesis that human immunodeficiency viruses have recently arisen from a diverse pool of nonhuman primate immunodeficiency viruses. Our understanding of the molecular phylogenetic relationships between primate and nonprimate lentiviruses is less certain, partly because key intermediate forms are still to be discovered. DNA and protein sequence comparisons reveal uncanny dissimilarities, as well as similarities, among the genetic sequences of these complex retroviruses, thereby giving rise to the notion that primate lentiviruses are participants in "fast-forward" evolution.

DNA, Viral↗
PubMed1992

Reversion of the hprt mutant clone SP5 by intrachromosomal recombination.

The spontaneous hprt mutant clone SP5, derived from V79 Chinese hamster cells, was shown to exhibit a duplication of approximately 2 kb, including exon 2 and its flanking intron sequences, inserted into the intron 1 sequence of the hprt gene. The most striking feature of SP5 is that this clone is quite unstable, demonstrating an extremely high spontaneous reversion frequency. Molecular analysis of 25 independent revertant clones of SP5 indicated that they arose after precise deletion of the duplicated fragment in the hprt gene. Reversion of SP5 could be induced by agents which damage DNA by different mechanisms, but there was no correlation with induction of the forward mutations. Based on these results, we suggest that intrachromosomal recombination must be responsible for the spontaneous reversion of SP5. Genetic recombination in somatic cells has been suggested to be involved in the multistep process of carcinogenesis. Since the ability to induce intrachromosomal recombination in yeast has been shown to be highly correlated with non-mutagenic as well as mutagenic carcinogens, it is of great interest to investigate similar systems in mammalian cells. The SP5 cell line may be unique for such a purpose, since this mutant clone contains an endogenic marker for studying the process of intrachromosomal recombination.

Animals↗
PubMed1997

Anatomy, pathophysiology, and prevention of senile enophthalmia and associated herniated lower eyelid fat pads.

We describe in detail the anatomy and function of the "Lockwood suspensory ligament" and the interrelated function of the orbital contents responsible for the intraorbital position of the eyeball and fat. With age, or because of genetic disposition, the eyeball descends, reducing the space between it and the floor of the orbit. This will inevitably cause forward projection of the extraconical orbital fat, creating herniated fat pads and resulting in enophthalmia. Based on the volume of the bony orbit and its contents, it is likely that relocating, rather then removing, herniated fat pads will greatly improve and prevent the enophthalmia of aging and give the globe a position and a projection of youth. Based on the results of surgery using the capsulopalpebral flap, it is likely that a descended Lockwood suspensory ligament, rather than a weakened orbital septum, is the cause of herniated fat pads and enophthalmia. We feel neither a weakened orbital septum nor an overabundance of orbital fat nor a shallow orbit is responsible for either of these conditions. We give a detailed description of how to raise the globe, preserve and relocate herniated fat pads, and manage and prevent enophthalmia and obtain a beautiful, youthful looking eye.

Adipose Tissue↗
PubMed2002

Initial arm muscle activation in a planar ballistic arm movement with varying external force directions: a simulation study.

It has been shown in previous research that the initial phase of EMG for a punching movement remained almost unchanged regardless of whether an external force was applied to the arm. The purpose of the present study was to explain this finding with the help of simulations. A two-dimensional model of the arm actuated by 6 Hill-type muscles was used to simulate a punching movement in the horizontal plane from a prescribed starting position with 90 elbow flexion. Input to the model was the stimulation of the muscles, and output were, among others, muscle forces and segmental accelerations. A genetic algorithm was used to determine the muscle onset times that minimized movement duration and targeting error. In a subsequent forward simulation, the optimized muscle onset times for an unloaded punching movement were superimposed on the isometric stimulation necessary to hold the arm in the starting position while an external force was applied to the arm. The resulting movement was only slightly different from the unloaded movement. It appeared that because of the low level of isometric muscle force prior to the movement, and the high level of stimulation during the movement, muscle force was increased at a rate that was almost independent of the prior force level. These results confirmed the suggestion that the initial phase of EMG in ballistic movements is more related to the rate of change of force than to the absolute force level. It is hypothesized that this may simplify the task of the nervous system in the choice of initial muscle activity in ballistic arm movements because no adjustments to varying external forces are required

Arm↗
PubMed2006

Detection of a paternally inherited fetal mutation in maternal plasma by the use of automated sequencing.

The discovery of circulating fetal DNA in maternal blood has been an encouraging step forward in the prenatal diagnostic field. It has opened up the possibility of development of a noninvasive method for the genetic analysis of the fetus. Many techniques have been applied to the study of this fetal DNA, but automated sequencing has been seldom used. The intention of this study was to use the automated sequencing technique for the detection of a paternally inherited fetal mutation in maternal plasma. Maternal plasma samples from a pregnant woman, whose husband had a mutation (Q134X) in the RP2 gene, which is located in the X-chromosome, were collected at two different gestational ages (10th and 19th week of gestation) in order to determine whether the paternally inherited fetal mutation could be detected by automated sequencing. Restriction analysis was also performed to confirm the results. The fetal mutation was clearly detected in the maternal plasma by the use of automated sequencing. The automated sequencing enables the possibility of analyzing fetal sequences, at a nucleotide level, in order to detect mutations or polymorphisms which are distinguishable from maternal sequences.

Base Sequence↗
PubMed1985

[Multiple gangrene of the digits in a female patient with liver cirrhosis, diabetes mellitus and cryoglobulinemia].

A case is described with multiple gangrene of the fingers of a female, aged 55, with confirmed cirrhosis of liver and diabetes mellitus. Cryoglobulins were established in the serum. A disturbed immunological balance was present and the family loading with collagenosis raises the problem of genetic moments of the disturbed homeostasis. The existing diabetic microangiopathy, histologically confirmed by biopsy of skin and subcutaneous tissue, brings forward the question of the complex pathogenesis of gangrene. At the end of her stay in the clinic, the patient developed cerebral apoplexy.

Brain Ischemia↗
PubMed2006

Diagnosing familial hypercholesterolaemia: the relevance of genetic testing.

AIMS: We assembled a cohort of patients with familial hypercholesterolaemia (FH) for both basic and clinical research. We used a set of established diagnostic criteria to define FH. Some put forward that a definite diagnosis of FH is made when a mutation in the LDL-receptor (LDLR) gene is identified. We therefore set out to determine in these patients whether patients with a DNA diagnosis would differ significantly from those diagnosed clinically. METHODS AND RESULTS: We randomly selected 4000 hypercholesterolaemic patients from the Dutch Lipid Clinic network database. Phenotypical data were acquired by reviewing medical records. After review of medical records, 2400 patients could be defined as having FH. An LDLR mutation was identified in 52.3% of these patients. Patients with and without an LDLR mutation demonstrated different clinical and laboratory characteristics. Low-density lipoprotein cholesterol was higher in patients with an LDLR mutation, whereas triglycerides were higher in patients without an LDLR mutation. The phenotypic differences between the groups remained even after stratification for the presence or absence of tendon xanthomas. CONCLUSION: Despite the use of stringent clinical criteria to define FH patients, two cohorts could be identified within our study population, namely those patients with and those without an LDLR mutation. Our findings suggest that among those without an LDLR mutation, patients with other causes of dyslipidaemia may be present. These observations underline the relevance of genetic testing in FH for clinical practice, for screening purposes, and for research involving these patients.

Adult↗
PubMedDate not supplied

The present status of the 'carrier hypothesis' for chemosensory recognition of genetic individuality.

The hypothesis that soluble MHC Class I molecules could act as the vector which transports MHC-specific odours from the blood into the urine was put forward some ten years ago (Singh, Brown & Roser, 1987). Here, I summarise new evidence in favour of the hypothesis. 'We propose that the ability of MHC Class I molecules to associate in a selective way with other small molecules could also be the mechanism by which a unique mixture of volatile, endogenous metabolites is transported by [soluble] class I MHC glycoproteins from the blood into the urine' (Singh, Brown & Roser, 1987).

Animals↗
PubMed1976

Evaluation and re-evaluation of genetic radiation hazards in man. I. Interspecific comparison of estimates of mutation rates.

A detailed presentation is made of the experimental data from the various systems used by Abrahamson et al. [2] to conclude that the per locus per rad (low LET) radiation-induced forward mutation rates in organisms, whose DNA content varies by a factor of about 1000, is proportional to genome size. Additional information pertinent in this context is also reviewed. It is emphasized that the mutation rates cited by Abrahamson et al. [2], although considered as pertaining to mutations at specific loci, actually derive from a broad variety of genetic end-points. It is argued that an initial (if not sufficient) condition for sound inter-specific mutation rate comparisions, covering a wide range of organisms and detecting systems of various sensitivities, requires a reasonalbly consistent biological definition of a specific locus mutation, namely, a transmissible intra-locus change. Granting the differences between systems in their resolving power to detect intragenic change, the data cited in this paper do not support the existence of a simple proportionality between radiotion-induced intra-locus mutation rate and genome size for the different species reviewed here. Furthermore, in Drosophila melanogaster, where individual salivary gland chromosome bands (that can differ greatly in DNA content) are usually associated with individual loci or at least distinct complementation groups, radiation-induced intra-locus mutation rates are not correlated with apparent differences in the DNA content of bands. This result is incompatible with the notion that most of the DNA in a band represents a radiation-mutable target capable of eliciting the kind of mutation observed in mutation rate experiments. All these considerations argue against the validity of the hypothesis of Abrahamson et al. [2] and their generalization that, for the evaluation of genetic radiation hazards in man, we can now "extrapolate from mutation rates obtained in lower organisms to man with greater confidence" on the basis of DNA content (italics are ours).

Animals↗
PubMed2026

Genetic interconnections between personality-related phenotypes and psychiatric disorders.

BACKGROUND: Personality-related phenotypes are genetically correlated with psychiatric disorders, but whether these relationships reflect shared genetic loci and differ across individual phenotypes remains unclear. We investigated their shared genetic architecture at the level of specific phenotype-disorder pairs. METHODS: We analyzed genome-wide association study summary statistics for 13 personality-related phenotypes and eight psychiatric disorders in populations of European ancestry. Genetic correlations were evaluated separately for 104 phenotype-disorder pairs using linkage disequilibrium score regression and high-definition likelihood. For pairs supported by both methods, MTAG and CPASSOC were applied separately to identify pleiotropic signals, followed by linkage disequilibrium clumping, Bayesian colocalization, gene prioritization, functional enrichment and bidirectional two-sample Mendelian randomization analyses. No composite personality or psychiatric-disorder phenotype was constructed. RESULTS: Among the 104 evaluated pairs, 77 showed significant positive genetic correlations in both analyses. Joint screening of MTAG and CPASSOC results identified pleiotropic signals in 61 pairs, comprising 1088 independent lead SNV-pair associations and 776 unique SNVs. Bayesian colocalization supported 351 signals across 42 pairs and 284 unique lead SNVs. MAGMA identified 1293 unique genes, of which 379 were prioritized by PoPS and 151 were further supported by SMR. These genes were enriched in brain tissues and biological processes involving nervous system development, synaptic organization and intercellular connectivity. Inverse-variance weighted Mendelian randomization identified 41 forward and 32 reverse associations after false-discovery-rate correction, including 21 pairs with bidirectional evidence. CONCLUSION: These item-resolved analyses identify widespread but heterogeneous genetic sharing between personality-related phenotypes and psychiatric disorders. The findings provide a pair-specific map of shared loci and prioritized genes, while the Mendelian randomization results should be interpreted cautiously because of residual heterogeneity and potential horizontal pleiotropy. Further validation in diverse populations and functional studies is required.

Colocalization↗
PubMed1994

Fluctuation test for two-stage mutations: application to gene amplification.

The determination of mutation rates is an important experimental procedure for characterizing mutation processes. The accepted method of determining mutation rates, the fluctuation test, was introduced by Luria and Delbrück in 1943. Since then it has been applied to various microorganisms and cells. The Luria-Delbrück test is based on a restrictive hypothesis of mutations being due to single irreversible events. However, some inherited changes in phenotype, like gene amplification, may be due to two or more genetic changes, some of which may be reversible. The Luria-Delbrück model of mutation was compared to other models which included reversibility and more than one mutation stage. The Luria-Delbrück model has been confirmed to be consistent with the original bacteriophage resistance data. However, for gene amplification this model gives incompatible estimates of mutation rates by the P0 and r methods. Relaxing the hypotheses of the single-stage models did not improve the fit. In contrast, a two-stage reversible model provided a fit. Analysis of gene amplification data by the two-stage reversible model provides new information, including estimates of rates for each of the two forward stages and of the reverse step.

Animals↗
PubMed2006

Utrastructural analysis of asthenozoospermic ejaculates in the era of assisted procreation.

OBJECTIVE: To define by transmission electron microscopy (TEM) analysis, the prevalence of sperm tail defects of genetic origin among men suffering for a reduced or absent sperm motility. DESIGN: A retrospective study. SETTING: Andrology Clinic, University of L'Aquila, Italy. PATIENT(S): The 120 ejaculates of infertile men with a forward motility (FM) < or =20% were compared to 200 ejaculates with a FM >20%. All ejaculates had a sperm vitality >50%. INTERVENTION(S): Some 25-50 tails were analyzed by TEM in each ejaculate. MAIN OUTCOME MEASURE(S): Receiver operating characteristics (ROC) analysis was applied by plotting the FM of cases with or without genetic tail defects detected by TEM. RESULT(S): The area under the ROC curve was 0.96 (95% confidence interval 0.92-0.98). The positive predictive accuracy for genetic tail defects in ejaculates with 0% FM was 46%. Three of 17 cases with genetic tail defects were classified as false negative when TEM analysis was restricted to ejaculates with 0% FM. A FM < or =7%, allowed the identification of all cases with genetic tail defects. CONCLUSION(S): The TEM analysis identifies sperm tail defects of genetic origin and should be restricted to ejaculates with severe asthenozoospermia (< or =7% motile sperm) and sperm vitality >50%.

Ejaculation↗
PubMed2005

Capillary electrophoresis single-strand conformation polymorphism analysis for monitoring soil bacteria.

The ability to effectively monitor a microbial community is necessary to design and implement remediation strategies for contaminated soil. Single-strand conformation polymorphism (SSCP), a technique which separates DNA fragments based on their sequence, was used to analyze amplified 16S rRNA gene fragments of 12 common soil bacteria. Separation was performed using capillary electrophoresis (CE), as opposed to other common gel techniques, to eliminate the need for band analysis on gel matrices. Four different universal bacterial primer sets were used for DNA amplification: 341-534, P11-P13, Er10-Er11, and Er14-Er15 corresponding to the V3, V8, V2, and V4 regions, respectively. The forward strand of each primer was labeled with 6-carboxy fluorescein fluorescent dye. Analyses were performed on the Applied Biosystems 310 genetic analyzer using GeneScan Analysis Software version 3.5. The best results were obtained using primer 341-534, in which 6 of the 12 bacteria could be distinguished. By combining primer sets 341-534 and Er10-Er11, all 12 of the bacteria could be separated, indicating various degrees of polymorphism within the selected primer regions. When performing simultaneous amplification and analysis of all 12 species some preferential amplification occurred, as not all peaks could be observed. However, SSCP profiles obtained for pure bacterial cultures show the potential of CE-SSCP for bacterial community analysis.

DNA, Bacterial↗