The CD14 receptor does not mediate entry of Mycobacterium tuberculosis into human mononuclear phagocytes

Prior reports have suggested that CD14 mediates uptake of Mycobacterium tuberculosis into porcine alveolar macrophages and human fetal microglia, but the contribution of CD14 to cell entry in human macrophages has not been studied. To address this question, we used flow cytometry to quantify uptake by human monocytes and alveolar macrophages of M. tuberculosis expressing green fluorescent protein. Neutralizing anti-CD14 antibodies did not affect bacillary uptake and the efficiency of bacillary entry was similar in THP-1 cells expressing low and high levels of CD14. However, most internalized bacteria were found in CD14+ but not in CD14- monocytes because M. tuberculosis infection upregulated CD14 expression. We conclude that: (1) CD14 does not mediate cellular entry by M. tuberculosis; (2) M. tuberculosis infection upregulates CD14 expression on mononuclear phagocytes, and this may facilitate the pathogen's capacity to modulate the immune response.     

NK cells regulate CD8+ T cell effector function in response to an intracellular pathogen

We studied the role of NK cells in regulating human CD8+ T cell effector function against mononuclear phagocytes infected with the intracellular pathogen Mycobacterium tuberculosis. Depletion of NK cells from PBMC of healthy tuberculin reactors reduced the frequency of M. tuberculosis-responsive CD8+IFN-gamma+ cells and decreased their capacity to lyse M. tuberculosis-infected monocytes. The frequency of CD8+ IFN-gamma+ cells was restored by soluble factors produced by activated NK cells and was dependent on IFN-gamma, IL-15, and IL-18. M. tuberculosis-activated NK cells produced IFN-gamma, activated NK cells stimulated infected monocytes to produce IL-15 and IL-18, and production of IL-15 and IL-18 were inhibited by anti-IFN-gamma. These findings suggest that NK cells maintain the frequency of M. tuberculosis-responsive CD8+IFN-gamma+ T cells by producing IFN-gamma, which elicits secretion of IL-15 and IL-18 by monocytes. These monokines in turn favor expansion of Tc1 CD8+ T cells. The capacity of NK cells to prime CD8+ T cells to lyse M. tuberculosis-infected target cells required cell-cell contact between NK cells and infected monocytes and depended on interactions between the CD40 ligand on NK cells and CD40 on infected monocytes. NK cells link the innate and the adaptive immune responses by optimizing the capacity of CD8+ T cells to produce IFN-gamma and to lyse infected cells, functions that are critical for protective immunity against M. tuberculosis and other intracellular pathogens.     

Effects of epistasis on phenotypic robustness in metabolic pathways

It is an open question whether phenomena such as phenotypic robustness to mutation evolve as adaptations or are simply an inherent property of genetic systems. As a case study, we examine this question with regard to dominance in metabolic physiology. Traditionally the conclusion that has been derived from Metabolic Control Analysis has been that dominance is an inevitable property of multi-enzyme systems and hence does not require an evolutionary explanation. This view is based on a mathematical result commonly referred to as the flux summation theorem. However it is shown here that for mutations involving finite changes (of any magnitude) in enzyme concentration, the flux summation theorem can only hold in a very restricted set of conditions. Using both analytical and simulation results we show that for finite changes, the summation theorem is only valid in cases where the relationship between genotype and phenotype is linear and devoid of non-linearities in the form of epistasis. Such an absence of epistasis is unlikely in metabolic systems. As an example, we show that epistasis can arise in scenarios where we assume generic non-linearities such as those caused by enzyme saturation. In such cases dominance levels can be modified by mutations that affect saturation levels. The implication is that dominance is not a necessary property of metabolic systems and that it can be subject to evolutionary modification.     

Uptake and Utilization of Double-Labeled High-Density Lipoprotein Sphingomyelin in Isolated Brain Capillaries of Adult Rats

Isolated rat brain capillaries were incubated in the presence of high-density lipoprotein (HDL) containing [stearic acid-14C, (methyl-3H)choline]sphingomyelin. This double-labeled sphingomyelin was taken up in a concentration-dependent manner. Cerebral capillary-associated sphingomyelin had a 3H/14C ratio close to that of the incubation medium, a result indicating uptake of sphingomyelin without prior hydrolysis. TLC of lipid extracted from capillaries showed that part of the sphingomyelin (up to 40%) was hydrolyzed in the brain capillaries to ceramide and free fatty acids. The hydrolysis was proportional to the amount of incorporated sphingomyelin and reached a plateau when the HDL sphingomyelin concentration in the medium was 237 nmol/ml. The results of "pulse-chase" experiments showed that the choline moiety of sphingomyelin was recovered in the incubation medium after the chase period and that there was no redistribution of liberated choline in phosphatidyl-choline of capillaries.     

A POPULATION GENETIC THEORY OF CANALIZATION

Canalization is the suppression of phenotypic variation. Depending on the causes of phenotypic variation, one speaks either of genetic or environmental canalization. Genetic canalization describes insensitivity of a character to mutations, and the insensitivity to environmental factors is called environmental canalization. Genetic canalization is of interest because it influences the availability of heritable phenotypic variation to natural selection, and is thus potentially important in determining the pattern of phenotypic evolution. In this paper a number of population genetic models are considered of a quantitative character under stabilizing selection. The main purpose of this study is to define the population genetic conditions and constraints for the evolution of canalization. Environmental canalization is modeled as genotype specific environmental variance. It is shown that stabilizing selection favors genes that decrease environmental variance of quantitative characters. However, the theoretical limit of zero environmental variance has never been observed. Of the many ways to explain this fact, two are addressed by our model. It is shown that a “canalization limit” is reached if canalizing effects of mutations are correlated with direct effects on the same character. This canalization limit is predicted to be independent of the strength of stabilizing selection, which is inconsistent with recent experimental data (Sterns et al. 1995). The second model assumes that the canalizing genes have deleterious pleiotropic effects. If these deleterious effects are of the same magnitude as all the other mutations affecting fitness very strong stabilizing selection is required to allow the evolution of environmental canalization. Genetic canalization is modeled as an influence on the average effect of mutations at a locus of other genes. It is found that the selection for genetic canalization critically depends on the amount of genetic variation present in the population. The more genetic variation, the stronger the selection for canalizing effects. All factors that increase genetic variation favor the evolution of genetic canalization (large population size, high mutation rate, large number of genes). If genetic variation is maintained by mutation-selection balance, strong stabilizing selection can inhibit the evolution of genetic canalization. Strong stabilizing selection eliminates genetic variation to a level where selection for canalization does not work anymore. It is predicted that the most important characters (in terms of fitness) are not necessarily the most canalized ones, if they are under very strong stabilizing selection (k > 0.2V_e). The rate of decrease of mutational variance V_m is found to be less than 10% of the initial V_m. From this result it is concluded that characters with typical mutational variances of about 10^–3 V_e are in a metastable state where further evolution of genetic canalization is too slow to be of importance at a microevolutionary time scale. The implications for the explanation of macroevolutionary patterns are discussed.     

Ghrelin Is Effective on Passive Avoidance Memory by Altering the Expression of NMDAR and HTR1a Genes in the Hippocampus of Male Wistar Rats

Background:Memory-dependent psychological behaviors have an important role in life. Memory strengthening in adulthood to prevent its defects in aging is a significant issue. The ghrelin endogenous hormone improves memory by targeting glutamatergic and serotonergic circuits. Also, citicoline, a memory strengthening drug in aging, is not recommended to adults due to its side effects. The current study aims to test that ghrelin treatment, like citicoline, would improve passive avoidance memory via expression of the genes encoding the N-methyl-D-aspartate receptor (NMDAR1) and the serotonin receptor 1A (HTR1α) involved in this process.Methods:Five groups of adult male rats received (1) saline (as control), (2) 0.5 mg/kg citicoline, or (3-5) 0.3, 1.5, and 3 nmol/μl ghrelin). The rats received the drugs via intra-hippocampal injection. Passive avoidance memory was determined using a shuttle box device. The latency to enter the dark chamber before (IL) and after (RL) injection and the total duration of the animal''s presence in the light compartment (TLC) were evaluated. Then, the gene expression rates of NMDAR1 and HTR1α were measured by the Real-Time PCR. Results:Ghrelin and citicoline had some similar and significant effects on passive avoidance memory, and both increased NMDAR1 and decreased HTR1α expression.Conclusion:Ghrelin, like citicoline, improves passive avoidance learning by altering the NMDAR1 and HTR1α expression in the hippocampusKey Words: Citicoline, Ghrelin, HTR1α, Intrahippocampal injection, NMDAR1, Passive Avoidance Memory     

Polyandry and the decrease of a selfish genetic element in a wild house mouse population

Despite deleterious effects on individuals, the t haplotype is a selfish genetic element present in many house mouse populations. By distorting the transmission ratio, +/t males transmit the t haplotype to up to 90% of their offspring. However, t/t individuals perish in utero. Theoretical models based on these properties predict a much higher t frequency than observed, leading to the t paradox. Here, we use empirical field data and theoretical approaches to investigate whether polyandry is a female counterstrategy against the negative fitness consequences of such distorters. We found a significant decrease of the t frequency over a period of 5.5 years that cannot be explained by the effect of transmission ratio distortion and recessive lethals, despite significantly higher life expectancy of +/t females compared to +/+ females. We quantified life-history data and homozygous and heterozygous fitness effects. Population subdivision and inbreeding were excluded as evolutionary forces influencing the t system. The possible influence of polyandry on the t system was then investigated by applying a stochastic model to this situation. Simulations show that polyandry can explain the observed t dynamics, making it a biologically plausible explanation for low t frequencies in natural populations in general.     

Alteration in Fatty Acid Composition of Adult Rat Brain Capillaries and Choroid Plexus Induced by a Diet Deficient in n-3 Fatty Acids: Slow Recovery After Substitution with a Nondeficient Diet

Wistar rats were fed for three generations with a semisynthetic diet containing either 1.5% sunflower oil (940 mg% of C18:2n-6, 6 mg% of C18:3n-3) or 1.9% soya oil (940 mg% of C18:2n-6, 130 mg% of C18:3n-3). At 60 days of age, the male offspring of the third generation were killed. The fatty acyl composition of isolated capillaries and choroid plexus was determined. The major changes noted in the fatty acid profile of isolated capillaries were a reduction (threefold) in the level of docosahexaenoic acid and, consequently, a fourfold increase in docosapentaenoic acid in sunflower oil-fed animals. The total percentage of polyunsaturated fatty acids was close to that in the soya oil-fed rats, but the ratio of n-3/n-6 fatty acids was reduced by threefold. In the choroid plexus, the C22:6n-3 content was also reduced, but by 2.6-fold, whereas the C22:5n-6 content was increased by 2.3-fold and the ratio of n-3/n-6 fatty acids was reduced by 2.4-fold. When the diet of sunflower oil-fed rats was replaced with a diet containing soya oil at 60 days of age, the recovery in content of n-6 and n-3 fatty acids started immediately after diet substitution; it progressed slowly to reach normal values after 2 months for C22:6n-5 and 2.5 months for C22:6n-3. The recovery in altered fatty acids of choroid plexus was also immediate and very fast. Recovery in content of C22:5n-6 and C22:6n-3 was complete by 46 days after diet substitution.     

Metabolic and endocrinologic complications in beta-thalassemia major: a multicenter study in Tehran

ABSTRACT : BACKGROUND : The combination of transfusion and chelation therapy has dramatically extended the life expectancy of thalassemic patients. The main objective of this study is to determine the prevalence of prominent thalassemia complications. METHODS : Two hundred twenty patients entered the study. Physicians collected demographic and anthropometric data and the history of therapies as well as menstrual histories. Patients have been examined to determine their pubertal status. Serum levels of 25(OH) D, calcium, phosphate, iPTH were measured. Thyroid function was assessed by T3, T4 and TSH. Zinc and copper in serum were determined by flame atomic absorption spectrophotometry. Bone mineral density (BMD) measurements at lumbar and femoral regions have been done using dual x-ray absorptiometry. The dietary calcium, zinc and copper intakes were estimated by food-frequency questionnaires. RESULTS : Short stature was seen in 39.3% of our patients. Hypogonadism was seen in 22.9% of boys and 12.2% of girls. Hypoparathyroidism and primary hypothyroidism was present in 7.6% and 7.7% of the patients. About 13 % of patients had more than one endocrine complication with mean serum ferritin of 1678 +/- 955 micrograms/lit. Prevalence of lumbar osteoporosis and osteopenia were 50.7% and 39.4%. Femoral osteoporosis and osteopenia were present in 10.8% and 36.9% of the patients. Lumbar BMD abnormalities were associated with duration of chelation therapy. Low serum zinc and copper was observed in 79.6% and 68% of the study population respectively. Serum zinc showed significant association with lumbar but not femoral BMD. In 37.2% of patients serum levels of 25(OH) D below 23 nmol/l were detected. CONCLUSION : High prevalence of complications among our thalassemics signifies the importance of more detailed studies along with therapeutic interventions.     

NMR structural studies reveal a novel protein fold for MerB, the organomercurial lyase involved in the bacterial mercury resistance system

Mercury resistant bacteria have developed a system of two enzymes (MerA and MerB), which allows them to efficiently detoxify both ionic and organomercurial compounds. The organomercurial lyase (MerB) catalyzes the protonolysis of the carbon-mercury bond resulting in the formation of ionic mercury and a reduced hydrocarbon. The ionic mercury [Hg(II)] is subsequently reduced to the less reactive elemental mercury [Hg(0)] by a specific mercuric reductase (MerA). To better understand MerB's unique enzymatic activity, we used nuclear magnetic resonance (NMR) spectroscopy to determine the structure of the free enzyme. MerB is characterized by a novel protein fold consisting of three noninteracting antiparallel beta-sheets surrounded by six alpha-helices. By comparing the NMR data of free MerB and the MerB/Hg/DTT complex, we identified a set of residues that likely define a Hg/DTT binding site. These residues cluster around two cysteines (C(96) and C(159)) that are crucial to MerB's catalytic activity. A detailed analysis of the structure revealed the presence of an extensive hydrophobic groove adjacent to this Hg/DTT binding site. This extensive hydrophobic groove has the potential to interact with the hydrocarbon moiety of a wide variety of substrates and may explain the broad substrate specificity of MerB.