Use of the Reverse Transcription-Polymerase Chain Reaction for the Detection of Pelargonium Flower Break Carmovirus

A polymerase chain reaction (PCR)-based assay was used to detect pelargonium flower break carmovirus (PFBV) in total RNA extractions made from infected Pelargonium plants. Extracts were reverse transcribed (RT) and the resultant cDNA was amplified by PCR, using oligonucleotide primers specific for 343, 510 and 832 base pair fragments of the RNA-dependent RNA polymerase gene of PFBV.
The specificity and sensitivity of RT-PCR were compared with the enzyme-linked immunosorbent assay (ELISA) for the detection of PFBV in Pelargonium tissues. The virus could be detected efficiently in high dilutions of sap from infected plants and at low concentrations of purified virus. Although ELISA is a powerful tool for virus detection, RT-PCR was over 1000 times more sensitive in detecting PFBV in leaf extracts of infected Pelargonium than was ELISA. The limit of detecting PFBV RNA by RT-PCR was 200 fg, compared with 200 pg of virus by ELISA.     

Modification of the erythrocyte membrane by a non-specific lipid transfer protein

The non-specific phospholipid transfer protein purified from bovine liver has been used to modify the phospholipid content and phospholipid composition of the membrane of intact human erythrocytes. Apart from an exchange of phosphatidylcholine between the red cell and PC-containing vesicles, the protein appeared to facilitate net transfer of phosphatidylcholine from the donor vesicles to the erythrocyte and sphingomyelin transfer in the opposite direction. Phosphatidylcholine transfer was accompanied by an equivalent transfer (on a molar basis) of cholesterol. An increase in phosphatidylcholine content in the erythrocyte membrane from 90 to 282 nmol per 100 microliters packed cells was observed. Phospholipase C treatment of modified cells showed that all of the phosphatidylcholine which was transferred to the erythrocyte was incorporated in the lipid bilayer. The nonspecific lipid transfer protein used here appeared to be a suitable tool to modify lipid content and composition of the erythrocyte membrane, and possible applications of this approach are discussed.     

Reversible (G0) and nonreadily reversible (Q) noncycling cells in human peripheral blood. Immunological, structural, and biological characterization

PHA-stimulated human lymphocytes (normal-resting-proliferating) at 0, 24, 48, 72, and 144 h were studied with Acridine Orange (AO) staining. By viable cell sorting, by subsequent subculturing, and by the use of biochemical, biophysical, and immunological assays, not only have the G0 resting and G1 (cycling) cell cycle phases been objectively characterized, but a separate subpopulation of quiescent cells that are functionally viable and deeply committed to nonproliferation, the Q cells, has been identified. Multiparameter cytofluorimetric analysis, methyl14C-thymidine incorporation, automated image analysis, and mitogen stimulation studies have shown that the "Q" cell, compared to the "G0" resting but easily recruitable cell, exhibits quite lower red and green AO emission, possesses 2c to 4c DNA content (rather than only 2c), has a higher average optical density, and is either nonrecruitable or recruitable-with-difficulty in PHA-stimulated lymphocyte cultures.     

Effects of fetal intravenous glucose challenge in normal and growth retarded fetuses

Fetal intravenous glucose challenge test (0.75 g/kg of estimated fetal weight) was performed at 26-33 weeks gestation in 9 patients undergoing fetal blood sampling (FBS) by ultrasound guided needling from the umbilical vein. The indication for FBS was rapid karyotyping for fetal malformations in 5 (control group) and severe intrauterine growth retardation in the remaining 4 (IUGR group). Fetal blood samples were taken before the glucose infusion and after 1, 3, 5, 10 and 15 min; glucose and insulin were assayed on each occasion and acid-base balance at 0 and 5 min. Basal fetal pO2, pH, glucose and insulin were lower in the IUGR group than in controls. Following the glucose challenge, fetal glucose levels were similar in the two groups, but in the IUGR group the latter part of the glucose curve was characterized by a slower and delayed return to basal levels. In control fetuses the insulin response following the glucose challenge peaked at 3 min while in IUGR no change in insulin concentration was detected. Fetal pO2 did not change in either group; the median change in fetal pH was significantly different between the two groups (controls: +0.01; IUGR: -0.04; P less than 0.05) and there was a significant correlation between basal pO2 and the change in fetal pH (r = 0.79) (P less than 0.02). These results support the concept of a low energy state in IUGR. Fetal glucose supplementation in IUGR is unlikely to be of benefit and may even exacerbate underlying acidosis.     

Predominance of bacteriophage SP82 over bacteriophage SP01 in mixed infections of Bacillus subtilis

In mixed infections with Bacillus subtilis phages SP82 and SP01, the SP82 genotype is predominant among the progeny. This predominance is determined by a specific region of the genome, the pos region, which apparently is located near genes 29 to 32 (by the SP01 numbering system). Recombination between SP82 and SP01 yields phage which have both the SP82 pos region and an SP01 mutation. This mutation then behaves in mixed infection as if it were part of an SP82 genome.     

Studies on the possible biological effects of 50 Hz electric and/or magnetic fields: Evaluation of some glycolytic enzymes,glycolytic flux,energy and oxido-reductive potentials in human erythrocytes exposed in vitro to power frequency fields

An attempt has been made to understand whether 50 Hz electric and magnetic fields (EMFs) are involved in producing bioeffects by exposing human erythrocytes in vitro. The study evaluated some key glycolytic enzymes, glucose consumption, lactate production, energy charge, 2,3-diphosphoglycerate, and reduced glutathione levels. all of which are biochemical parameters significant to erythrocyte function. Cells exposed to individual or superimposed EMFs have not shown any significant difference compared with the controls. © 1993 Wiley-Liss. Inc.     

A Dual Strategy to Cope with High Light in Chlamydomonas reinhardtii

Absorption of light in excess of the capacity for photosynthetic electron transport is damaging to photosynthetic organisms. Several mechanisms exist to avoid photodamage, which are collectively referred to as nonphotochemical quenching. This term comprises at least two major processes. State transitions (qT) represent changes in the relative antenna sizes of photosystems II and I. High energy quenching (qE) is the increased thermal dissipation of light energy triggered by lumen acidification. To investigate the respective roles of qE and qT in photoprotection, a mutant (npq4 stt7-9) was generated in Chlamydomonas reinhardtii by crossing the state transition–deficient mutant (stt7-9) with a strain having a largely reduced qE capacity (npq4). The comparative phenotypic analysis of the wild type, single mutants, and double mutants reveals that both state transitions and qE are induced by high light. Moreover, the double mutant exhibits an increased photosensitivity with respect to the single mutants and the wild type. Therefore, we suggest that besides qE, state transitions also play a photoprotective role during high light acclimation of the cells, most likely by decreasing hydrogen peroxide production. These results are discussed in terms of the relative photoprotective benefit related to thermal dissipation of excess light and/or to the physical displacement of antennas from photosystem II.     

Mixing Nulliparous and Multiparous Women in Randomised Controlled Trials of Preeclampsia Prevention Is Debatable: Evidence from a Systematic Review

Background

Nulliparity is a major risk factor of preeclampsia investigated in numerous trials of its prevention.

Objective

We aimed to assess whether these trials considered nulliparity in subject selection or analysis of results.

Search Strategy

01 April 2013 search of MEDLINE via PubMed, EMBASE and the Cochrane Library. 01 April 2013 search of trials registered in Clinicaltrials.gov.

Selection Criteria

Randomised controlled trials and metaanalyses of preeclampsia prevention with no restriction to period of publication or language. Metaanalyses were selected to fully identify relevant trials.

Data Collection and Analysis

One reader appraised each selected article/registered protocol using a pretested, standardized data abstraction form developed in a pilot test. For each article, he recorded whether both nulliparous and multiparous were included and, in case of mixed populations, whether randomisation was stratified, and whether subgroup analyses had been reported. For registered protocols, he only assessed whether it was planned to include mixed populations.

Main Results

88 randomised controlled trials were identified, representing 83,396 included women. In 58 of the 88 articles identified (65.9%), preeclampsia was the primary outcome. In 31 of these (53.4%), the investigation combined nulliparous and multiparous women; only two reports in 31 (6.5%) stated that randomisation was stratified on parity and only four (12.9%) described a subgroup analysis by parity. Of the 30 registered trials, 20 (66.6%) planned to include both nulliparous and multiparous women.

Conclusion

Parity is largely ignored in randomised controlled trials of preeclampsia prevention, which raises difficulties in interpreting the results.     

Implanting Glass Spinal Cord Windows in Adult Mice with Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) in adult rodents is the standard experimental model for studying autonomic demyelinating diseases such as multiple sclerosis. Here we present a low-cost and reproducible glass window implantation protocol that is suitable for intravital microscopy and studying the dynamics of spinal cord cytoarchitecture with subcellular resolution in live adult mice with EAE. Briefly, we surgically expose the vertebrae T12-L2 and construct a chamber around the exposed vertebrae using a combination of cyanoacrylate and dental cement. A laminectomy is performed from T13 to L1, and a thin layer of transparent silicone elastomer is applied to the dorsal surface of the exposed spinal cord. A modified glass cover slip is implanted over the exposed spinal cord taking care that the glass does not directly contact the spinal cord. To reduce the infiltration of inflammatory cells between the window and spinal cord, anti-inflammatory treatment is administered every 2 days (as recommended by ethics committee) for the first 10 days after implantation. EAE is induced only 2-3 weeks after the cessation of anti-inflammatory treatment.Using this approach we successfully induced EAE in 87% of animals with implanted windows and, using Thy1-CFP-23 mice (blue axons in dorsal spinal cord), quantified axonal loss throughout EAE progression. Taken together, this protocol may be useful for studying the recruitment of various cell populations as well as their interaction dynamics, with subcellular resolution and for extended periods of time. This intravital imaging modality represents a valuable tool for developing therapeutic strategies to treat autoimmune demyelinating diseases such as EAE.