Hormonal control of reversible translocation of perilipin B to the plasma membrane in primary human adipocytes

In adipocytes, perilipin coats and protects the central lipid droplet, which stores triacylglycerol. Alternative mRNA splicing gives rise to perilipin A and B. Hormones such as catecholamines and insulin regulate triacylglycerol metabolism through reversible serine phosphorylation of perilipin A. It was recently shown that perilipin was also located in triacylglycerol-synthesizing caveolae of the plasma membrane. We now report that perilipin at the plasma membrane of primary human adipocytes was phosphorylated on a cluster of threonine residues (299, 301, and 306) within an acidic domain that forms part of the lipid targeting domain. Perilipin B comprised <10% of total perilipin but was the major isoform associated with the plasma membrane of human adipocytes. This association was controlled by insulin and catecholamine: perilipin B was specifically depleted from the plasma membrane in response to the catecholamine isoproterenol, while insulin increased the amount of threonine phosphorylated perilipin at the plasma membrane. The reversible translocation of perilipin B to and from the plasma membrane in response to insulin and isoproterenol, respectively, suggests a specific function for perilipin B to protect newly synthesized triacylglycerol in the plasma membrane.     

Validation of an Instrument to Measure Older Adults' Expectations Regarding Movement (ERM)

Background

Many individuals with Parkinson''s disease are not diagnosed and treated. Attitudes about aging and related help-seeking may affect the timely diagnosis of Parkinson''s disease. Our objectives were to develop measures of older adults'' expectations regarding movement with aging, specifically related to parkinsonism, and their beliefs about seeking healthcare for the diagnosis and treatment of parkinsonism.

Methods

We established content and face validity from interviews with experts, review of the literature, and pre-testing with key informants. Two 9-item instruments resulted: Expectations Regarding Movement (ERM) and Healthcare Seeking Beliefs for parkinsonism (HSB). These instruments were administered to 210 older adults at senior centers to investigate internal consistency and construct validity.

Results

192 (91%) of the older adults completed more than 90% of the survey. The mean age was 76; 17 (9%) reported parkinsonism. Both scales demonstrated good internal consistency (α = 0.90). Factor analysis supported construct validity of the ERM and HSB scores. Older age, lower education, worse self-reported health and African American race each were associated with lower ERM scores, but not HSB scores.

Conclusion

The ERM, a brief measure of expectations regarding movement with aging, shows reliability and validity. This scale may be useful in identifying older adults at increased risk for under-identification of Parkinson''s disease. Further work is needed to measure healthcare seeking for parkinsonism.     

Induction of cell death in rat brain by a gliotoxic factor from cerebrospinal fluid in multiple sclerosis.

The cerebrospinal fluid (CSF) of multiple sclerosis (MS) patients contains a 17 kDa glycoproteic factor with gliotoxic properties in vitro. In order to study the physiopathological role of this gliotoxic factor in vivo, we have injected a partially purified preparation and appropriate controls in rat CSF and investigated whether it induces cell death in the rat central nervous system (CNS), 10 days and 3 months after injection. We used the TUNEL assay in association with specific immunohistochemistry to characterize dying cells in the gliotoxic factor- treated rat CNS. At 10 days post-injection, TUNEL-positive cells were observed in the whole rat CNS. They were particularly numerous in the choroid plexus, ependymal epithelium, cerebral white matter, cerebral vascular endothelium, arachnoid spaces and less frequent in the gray matter of brain and spinal cord. The predominant type of TUNEL-positive cells observed at 10 days post-injection was astrocytes, in white matter, gray matter, occasionnally around damaged endothelial cells in periventricular and subpial spaces. Other TUNEL-positive cells were identified as oligodendrocytes by an oligodendrocyte specific RIP immunostaining, at 10 days post-treatment with the gliotoxic factor. Interestingly, demyelination and death of oligodendrocytes were more important 3 months post-injection: TUNEL-RIP positive oligodendrocytes were generally associated with multifocal demyelinating areas. Clearly, the 17 kDa gliotoxic factor injection in rat CSF triggers demyelination and may be used as a new animal model for MS. Also, our results suggest a new possible scenario for MS pathogenesis: death of oligodendrocytes and astrocytes, stimulated by the MS gliotoxic factor causes the breakdown of the blood-brain barrier (BBB) and the demyelinating cascade.     

Cryopreservation of sour orange (Citrus aurantium L.) shoot tips

Summary The objective of this study was to establish a cryopreservation protocol for sour orange (Citrus aurantium L.). Cryopreservation was carried out via encapsulation-dehydration, vitrification, and encapsulation-vitrification on shoot tips excised from in vitro cultures. Results indicated that a maximum of 83% survival and 47% regrowth of encapsulated-dehydrated and cryopreserved shoot tips was obtained with 0.5M sucrose in the preculture medium and further dehydration for 6 h to attain 18% moisture content. Dehydration of encapsulated shoot tips with silica gel for 2h resulted in 93% survival but only 37% regrowth of cryopreserved shoot tips. After preculturing with 0.5M sucrose, 80% of the vitrified cryopreserved shoots survived when 2M sucrose plus 10% dimethyl sulfoxide (DMSO) was used as a cryoprotectant for 20 min at 25°C. Survival and regrowth of vitrified cryopreserved shoot tips were 67% and 43%, respectively, when 0.4M sucrose plus 2M glycerol was used as a loading solution followed by application of 100% plant vitrification solution (PVS2) for 20 min. Increased duration of exposure to the loading solution up to 60 min increased survival (83%) and regrowth (47%) of cryopreserved shoot tips. With encapsulation-vitrification, dehydration with 100% PVS2 for 2 or 3 h at 0°C resulted in 50 or 57% survival and 30 or 40% regrowth, respectively, of cryopreserved shoot tips.     

Bee venom ameliorates cardiac dysfunction in diabetic hyperlipidemic rats

High levels of blood glucose and lipids are well-known risk factors for heart diseases. Bee venom is a natural product that has a potent hypoglycemic, hypolipidemic, anti-inflammatory, and antioxidant effects. The current study aimed to determine the bee venom effects on cardiac dysfunction compared to combined therapy of metformin and atorvastatin in diabetic hyperlipidemic rats. The median lethal dose of bee venom was estimated, and then 50 adult male albino rats were categorized into five groups. One group was fed a standard diet and served as a negative control, while the other groups were given nicotinamide and streptozotocin injections to induce type 2 diabetes. After confirming diabetes, the rats were fed a high-fat diet for four weeks. The four groups were divided as follows: one group served as a positive control, whereas the other three groups were treated with bee venom (0.5 mg/kg), bee venom (1.23 mg/kg), and combined therapy of metformin (60 mg/kg) and atorvastatin (10 mg/kg), respectively, for four weeks. Upon termination of the experiment, blood samples and heart tissue were obtained. Administration of bee venom using both doses (0.5 and 1.23 mg/kg) and combined therapy of metformin and atorvastatin revealed a significant decrease in the concentrations of glucose, total cholesterol, triacylglycerol, low-density lipoprotein cholesterol, very low-density lipoprotein cholesterol, troponin I, creatine kinase, and lactate dehydrogenase activities. Moreover, a significant decrease had been detedcted in malondialdehyde, nuclear factor-kappa-β levels, and relative mRNA expression of vascular cell adhesion molecule-1 and galectin-3 in heart tissue compared to the positive control (P < 0.0001). Furthermore, there was a significant increase in bodyweight levels of insulin, high-density lipoprotein cholesterol, and total antioxidant capacity in heart tissue compared to the positive control (P < 0.0001). The results indicate that bee venom can ameliorate cardiac dysfunction through attenuating oxidative stress and downregulating the NF-κβ signaling pathway.     

Visualization of body thermoregulation by infrared imaging

This paper analyzes two mechanisms applied on the human body in order to study the thermoregulatory system according to heat generation and heat loss. Two approaches are presented. The first approach is based on plethysmography, where an armband is placed on the forearm in order to modulate the blood flow. The second approach uses a cold stimulation. The visualization is achieved using infrared imaging devices. The resulting images reveal a temperature balance between the stimulated and the non-stimulated hands. The thermal behavior and typical thermographic recordings on each subject are discussed and analyzed in response to different stimulations.     

Independent evolution of functional Pm3 resistance genes in wild tetraploid wheat and domesticated bread wheat

The Pm3 alleles of cultivated bread wheat confer gene for gene resistance to the powdery mildew fungus. They represent a particular case of plant disease resistance gene evolution, because of their recent origin and possible evolution after the formation of hexaploid wheat. The Pm3 locus is conserved in tetraploid wheat, thereby allowing the comparative evolutionary study of the same resistance locus in a domesticated species and in one of its wild ancestors. We have identified 61 Pm3 allelic sequences from wild and domesticated tetraploid wheat subspecies. The Pm3 sequences corresponded to 24 different haplotypes. They showed low sequence diversity, differing by only a few polymorphic sequence blocks that were further reshuffled between alleles by gene conversion and recombination. Polymorphic sequence blocks are different from the blocks found in functional Pm3 alleles of hexaploid wheat, indicating an independent evolution of the Pm3 loci in the two species. A new functional gene was identified in a wild wheat accession from Syria. This gene, Pm3k , conferred intermediate race-specific resistance to powdery mildew, and consists of a mosaic of gene segments derived from non-functional alleles. This demonstrates that Pm3 -based resistance is not very frequent in wild tetraploid wheat, and that the evolution of functional resistance genes occurred independently in wild tetraploid and bread wheat. The Pm3 sequence variability and geographic distribution indicated that diversity was higher in wild emmer wheat from the Levant area, compared with the accessions from Turkey. Further screens for Pm3 functional genes in wild wheat should therefore focus on accessions from the Levant region.     

Phytoaccumulation of zinc by the aquatic plant, Lemna gibba L.

The uptake of zinc (Zn) by the duckweed Lemna gibba L., native to the north-east region of Algeria, was investigated in quarter Coïc solutions enriched with 6.0, 10.0, 14.0 and 18.0 mg l⁻¹ of Zn supplied as zinc sulphate (ZnSO₄). Zinc concentrations were measured in the water daily and in duckweed biomass at the end of the experiments. These results showed that under experimental conditions (pH = 6.0 ± 0.1, T = 21 ± 1 °C, photoperiod = 12 h/j), L. gibba was able to accumulate in its biomass 4.23; 15.62; 23.88 and 25.81 mg g⁻¹ DM, respectively for the four initial concentrations selected. At these concentrations, the metal removed percentages were 61–71%. The mass balance performed on the system showed that about 49–68% of Zn (depending on the initial concentration in water) was removed by precipitation as zinc phosphate. The results showed that this aquatic plant can be successfully used for Zn removal.     

Biophysical Characterization of <Emphasis Type="Italic">β</Emphasis>-Thalassemic Red Blood Cells

Thalassemia is the world’s most common hereditary disease; therefore, more interest has been devoted for the development of the screening procedure of this disease. In β-thalassemia major, the subject of the current study, impaired biosynthesis of beta-globin leads to accumulation of unpaired alpha-globin chain. The objective of the present study, was to examine many of the biophysical properties of β-thalassemia major red blood cells (RBCs) and to study the possibility of use of any of them as a preliminary screening tool for β-thalassemia. The percentage of normal hemolysis, osmotic fragility test, turbidity test, rheological properties, and dielectric properties, were studied in 20 regularly blood transfused thalassemia major patients who were under chelation therapy and their status were compared with those of 10 healthy subjects. There was an increase in the percentage of hemolysis for β-thalassemia by 114.6% compared to the normal RBCs. The fragility curve for β-thalassemia RBCs showed a shift toward lower NaCl concentration compared to the normal curve. The average osmotic fragility (H ₅₀: the NaCl concentration producing 50% homolysis) for β-thalassemia was found to be 3.21 ± 0.67 g/l, whereas for normal RBCs it was 5.5 ± 0.31 g/l. The turbidity curve of the β-thalassemic RBCs showed a shift toward higher detergent concentration of the normal curve, with higher value for the average membrane solubilization (S ₅₀). The viscosity value of whole blood β-thalassemia was found to be 3.916 ± 0.56 cp whereas for normal blood was 2.516 ± 0.36 cp. The relative permittivity, dielectric loss, and AC conductivity of RBCs decreased significantly compared to normal samples. This could be attributed to the loss of the insulating properties of the membrane and loss of its surface charge of thalassemic RBCs. As can be noticed, several factors showed clear difference between thalassemic and normal blood samples. Some of these parameters could be measured immediately after sample withdrawal and require short time to perform the measurements. This offers the advantages of being effective, low cost, and fast techniques, therefore, we suggest that these techniques could be applied for β-thalassemia major screening purposes.     

Infection of CD127+ (interleukin-7 receptor+) CD4+ cells and overexpression of CTLA-4 are linked to loss of antigen-specific CD4 T cells during primary human immunodeficiency virus type 1 infection

We recently found that human immunodeficiency virus (HIV)-specific CD4+ T cells express coreceptor CCR5 and activation antigen CD38 during early primary HIV-1 infection (PHI) but then rapidly disappear from the circulation. This cell loss may be due to susceptibility to infection with HIV-1 but could also be due to inappropriate apoptosis, an expansion of T regulatory cells, trafficking out of the circulation, or dysfunction. We purified CD38+++CD4+ T cells from peripheral blood mononuclear cells, measured their level of HIV-1 DNA by PCR, and found that about 10% of this population was infected. However, a small subset of HIV-specific CD4+) T cells also expressed CD127, a marker of long-term memory cells. Purified CD127+CD4+ lymphocytes contained fivefold more copies of HIV-1 DNA per cell than did CD127-negative CD4+ cells, suggesting preferential infection of long-term memory cells. We observed no apoptosis of antigen-specific CD4+ T cells in vitro and only a small increase in CD45RO+CD25+CD127dimCD4+ T regulatory cells during PHI. However, 40% of CCR5+CD38+++ CD4+ T cells expressed gut-homing integrins, suggesting trafficking through gut-associated lymphoid tissue (GALT). Furthermore, 80% of HIV-specific CD4+ T cells expressed high levels of the negative regulator CTLA-4 in response to antigen stimulation in vitro, which was probably contributing to their inability to produce interleukin-2 and proliferate. Taken together, the loss of HIV-specific CD4+ T cells is associated with a combination of an infection of CCR5+ CD127+ memory CD4+ T cells, possibly in GALT, and a high expression of the inhibitory receptor CTLA-4.