Transitions between distinct compaction regimes in complexes of multivalent cationic lipids and DNA

Cationic lipids (CLs) have found widespread use as nonviral gene carriers (vectors), including applications in clinical trials of gene therapy. However, their observed transfection efficiencies (TEs) are inferior to those of viral vectors, providing a strong incentive for a detailed understanding of CL-DNA complex behavior. In recent systematic studies employing monovalent as well as newly synthesized multivalent lipids (MVLs), the membrane cationic charge density has been identified as a key parameter governing the TE of lamellar CL-DNA complexes. In this work, we use x-ray scattering and molecular simulations to investigate the structural properties of complexes containing MVLs. At low mole fraction of neutral lipids (NLs), Φ_NL, the complexes show dramatic DNA compaction, down to essentially close-packed DNA arrays with a DNA interaxial spacing d_DNA = 25 Å. A gradual increase in Φ_NL does not lead to a continuous increase in d_DNA as observed for DNA complexes of monovalent CLs. Instead, distinct spacing regimes exist, with sharp transitions between the regimes. Three packing states have been identified: 1), close packed, 2), condensed, but not close packed, with d_DNA = 27-28 Å, and 3), an expanded state, where d_DNA increases gradually with Φ_NL. Based on our experimental and computational results, we conclude that the DNA condensation is mediated by the multivalent cationic lipids, which assemble between the negatively charged DNA rods. Quite remarkably, the computational results show that the less tightly packed structure in regime 2 is thermodynamically more stable than the close packed structure in regime 1. Accordingly, the constant DNA spacing observed in regime 2 is attributed to lateral phase coexistence between this stable CL-DNA complex and neutral membranes. This finding may explain the reduced TE measured for such complexes: transfection involves endosomal escape and disassembly of the complex, and these processes are inhibited by the high thermodynamic stability. Our results, which demonstrate the existence of an inverse correlation between the stability and transfection activity of lamellar CL-DNA complexes are, therefore, consistent with a recently proposed model of cellular entry.     

Loss of kindlin-1, a human homolog of the Caenorhabditis elegans actin-extracellular-matrix linker protein UNC-112, causes Kindler syndrome

Kindler syndrome is an autosomal recessive disorder characterized by neonatal blistering, sun sensitivity, atrophy, abnormal pigmentation, and fragility of the skin. Linkage and homozygosity analysis in an isolated Panamanian cohort and in additional inbred families mapped the gene to 20p12.3. Loss-of-function mutations were identified in the FLJ20116 gene (renamed “KIND1” [encoding kindlin-1]). Kindlin-1 is a human homolog of the Caenorhabditis elegans protein UNC-112, a membrane-associated structural/signaling protein that has been implicated in linking the actin cytoskeleton to the extracellular matrix (ECM). Thus, Kindler syndrome is, to our knowledge, the first skin fragility disorder caused by a defect in actin-ECM linkage, rather than keratin-ECM linkage.     

Hospitalized dogs recovery from naturally occurring heatstroke; does serum heat shock protein 72 can provide prognostic biomarker?

Heatstroke is a serious illness in dogs characterized by core temperatures above 41 °C with central nervous system dysfunction. Experimental heatstroke models have tried to correlate biomarker levels with the severity of the syndrome. Serum heat shock protein (eHSP70) levels were recently evaluated as a biomarker of heat tolerance and acclimation, their role as a marker of heatstroke is inconclusive. Here, we monitored eHSP70 levels in correlation with systemic biomarkers in 30 naturally occurring canine heatstroke cases. Thirty dogs diagnosed with environmental (33 %) or exertional (66 %) heatstroke admitted to hospital (0–14 h post-injury) were tested for biomarkers of organ damage and coagulation parameters. eHSP70 levels were measured upon admission and 4, 12, and 24 h later (T1, T2, and T3, respectively). No differences were found between exertional and environmental heatstroke cases. The eHSP profile demonstrated an inverted bell shape, with the lowest levels at the 12 h time point. A positive correlation between eHSP70, lactate, and aPPT was also noted at T2 in all the dogs in the study. Twenty-four h after presentation, eHSP70 levels returned to those measured upon admission, this change was only significant in the survivors. The obtained results suggest that eHSP72 level profile may be predictive of survival.     

Effects of Modafinil on Clonic Seizure Threshold Induced by Pentylenetetrazole in Mice: Involvement of Glutamate,Nitric oxide,GABA, and Serotonin Pathways

Epilepsy is the third most common chronic brain disorder. Modafinil is an awakening agent approved for narcolepsy. In addition to its clinical uses some reports revealed that modafinil was associated with some alterations in seizure threshold. The purpose of this study was to clarify the effect of acute administration of modafinil in clonic seizure threshold (CST) induced by pentylenetetrazole in mice and the involvement of glutamate, nitric oxide, gamma amino butyric acid (GABA), and serotonin systems in this feature. Modafinil at 80 and 150 mg/kg showed anti- and pro-convulsant effects respectively and expressed maximum anti- and pro-convulsant activities at 30 min after injection. Both modulatory effects were blunted by pretreatment of l-NAME [nonspecific nitric oxide synthase (NOS) inhibitor; 10 mg/kg, i.p.], 7-nitroindazole (a neuronal NOS inhibitor; 40 mg/kg, i.p.), and aminoguanidine (an inducible NOS inhibitor; 50 mg/kg, i.p.). Injection of the NOS precursor l-arginine (60 mg/kg, i.p.) before modafinil did not change the anti-convulsant effect, while thoroughly reversed the pro-convulsant one. Our experiments displayed that administration of diazepam (a GABA_A receptor agonist; 0.02 mg/kg, i.p.) and MK-801 (a NMDA receptor antagonist; 0.05 mg/kg, i.p.) before different doses of modafinil significantly increased CST. Finally, pretreatment of citalopram (a selective serotonin reuptake inhibitor; 0.1 mg/kg, i.p.) did not modify the convulsant activities of modafinil. Therefore, nitric oxide system may mediate anti-convulsant activity, while glutamate, nitric oxide, and GABA pathways may involve in pro-convulsant property. Serotonin receptors have no role on convulsant effects of modafinil.     

Systematic relationships within the Microtus arvalis (Rodentia: Cricetidae) group in Iran,inferred from cytogenetic analyses

The distribution of C-heterochromatin and nucleolar organizer regions (NORs) was studied in three species of voles of the Microtus arvalis group in Iran: M. mystacinus, M. kermanensis, and M. transcaspicus. The C-banding pattern and NORs distribution were similar in M. mystacinus and M. kermanensis suggesting taxonomic proximity of these two species. At the same time, the karyotypes of M. mystacinus from Iran were different in C-banding pattern from the complements of conspecific 54-chromosome voles from Europe and other regions of Asia. The most distinct difference was in size of the distal C-positive block of heterochromatin on the X chromosome. In this respect M. mystacinus from Iran and M. kermanensis resembled M. transcaspicus. Small size of the distal C-positive heterochromatic block may be ancestral whereas larger size is derived. The X chromosome of M. transcaspicus can be derived from that of M. mystacinus and M. kermanensis by a large inversion or centromeric shift.     

Attack, escape and predation rates of larvae of two aphidophagous ladybirds during conspecific and heterospecific interactions

The attack, escape and predation rates for larvae of aphidophagous ladybird Propylea dissecta (Mulsant) and Coccinella transversalis Fabricius were quantified as a potential mechanism leading to the differences in the incidence of cannibalism and intraguild predation. These rates were compared at four larval instars within and between the species. The attack rates of larvae of C. transversalis were significantly higher than those of P. dissecta towards conspecific and heterospecific victims. For both species, third instars exhibited maximum tendency to attack. Escape rates in C. transversalis were higher than P. dissecta. In P. dissecta, the second instars made a greater number of escapes than other conspecific instars after being attacked by same stage cannibal or heterospecific predator. In P. dissecta, first instars suffered maximum mortality due to cannibalism and intraguild predation by conspecifics and heterospecifics of the same and older developmental stage. No larvae of C. transversalis were eaten by P. dissecta of the same stage. These results suggest that the larvae of P. dissecta were more often potential cannibals than intraguild predators, while the reverse was the case in C. transversalis. Based on this finding, it could be predicted that in patchy prey habitats, high rates of larval cannibalism in P. dissecta would occur with a high risk of cannibalism of first instars. Larvae of C. transversalis would respond as intraguild predators, while those of P. dissecta as intraguild prey. The greater size and walking activity of C. transversalis could be possible reason for this tendency.     

Are developing countries prepared to face Ebola-like outbreaks?

<正>Ebola virus disease has caused havoc in West Africa,with 11,162deaths and more than 27,181 cases(as of May 31,2015)being reported since the virus emerged in early2014 in Guinea.The maximum number of cases has been reported in Sierra Leone(12,827),while most of the reported deaths have occurred in Liberia(4,806),according to the Center for Disease Control and     

Severe retinitis pigmentosa mapped to 4p15 and associated with a novel mutation in the PROM1 gene

Mutation in the PROM1 gene previously has been identified in one family with retinal degeneration for which neither ERG recordings nor detailed information about visual impairment is available. A large family with multiple individuals affected by retinal degeneration was ascertained in the Punjab province of Pakistan. The visual acuity of all affected patients in the family was severely compromised beginning in early childhood. The retinal disease in this family is a severe form of retinitis pigmentosa (RP) accompanied by macular degeneration. Fundus changes advanced with age. Choriocapillaris atrophy and posterior RPE atrophy were obvious allowing visualization of the large choroidal vessels in patients over 40 years of age. Rod and cone responses on ERG recordings were extinguished in patient’s teens. A genome-wide scan mapped the disease to a 34.7 cM region of chromosome 4p14–p16 between D4S1599 and D4S405. A maximum lod score of 3.96 with D4S403 and D4S391 is seen at θ = 0. Sequence analysis of PROM1 located in the linkage interval identified a c.1726C>T homozygous transition in exon 15: resulting in p.Gln576X in the translated protein. This mutation is found in a homozygous state in all six affected individuals and was heterozygous in five of the six unaffected family members examined. The mutation was not detected in 192 chromosomes of unrelated control individuals of the same ethnicity and from the same region. This delineates the phenotypic characteristics of retinopathy caused by mutations in PROM1. Qingjiong Zhang, Fareeha Zulfiqar, Xueshan Xiao, Sheikh Riazuddin and J. Fielding Hejtmancik contributed equally.     

Dephosphorylation of rabbit skeletal muscle glycogen synthase (phosphorylated by cyclic AMP-independent synthase kinase 1) by phosphatases

Phosphorylation of rabbit skeletal muscle glycogen synthase by cyclic AMP-independent synthase kinase 1 results in the incorporation of 4 mol of PO4/subunit. Incubation of the phosphorylated synthase with rabbit muscle phosphoprotein phosphatase brings about the hydrolysis of phosphates from all four major tryptic peptides and an increase in the synthase activity ratio from 0.01 to 0.85. Incubation of the phosphorylated synthase with calf intestinal alkaline phosphatase brings about the preferential hydrolysis of phosphates from three of the four major tryptic peptides and a slight increase in the four major tryptic peptides and a slight increase in the synthase activity ratio from 0.01 to 0.1. The phosphorylation site which is resistant to hydrolysis by calf intestinal alkaline phosphatase can be dephosphorylated by subsequent incubation with rabbit muscle phosphoprotein phosphatase. This dephosphorylation is accompanied by an increase in the synthase activity ratio to approximately 0.9. Measurements of the changes in the kinetic properties of the synthase samples dephosphorylated by alkaline phosphatase reveal that the phosphorylation sites susceptible to hydrolysis by alkaline phosphatase mainly affect the binding of glucose-6-P to the synthase. Comparison of the kinetic properties of the synthase samples dephosphorylated by alkaline phosphatase and by phosphoprotein phosphatase we find that the phosphorylation site resistant to hydrolysis by alkaline phosphatase affects both the binding of UDP-glucose and glucose-6-P to the synthase.