Brain barriers: Crosstalk between complex tight junctions and adherens junctions

Unique intercellular junctional complexes between the central nervous system (CNS) microvascular endothelial cells and the choroid plexus epithelial cells form the endothelial blood-brain barrier (BBB) and the epithelial blood-cerebrospinal fluid barrier (BCSFB), respectively. These barriers inhibit paracellular diffusion, thereby protecting the CNS from fluctuations in the blood. Studies of brain barrier integrity during development, normal physiology, and disease have focused on BBB and BCSFB tight junctions but not the corresponding endothelial and epithelial adherens junctions. The crosstalk between adherens junctions and tight junctions in maintaining barrier integrity is an understudied area that may represent a promising target for influencing brain barrier function.

Mutations in SDHD lead to autosomal recessive encephalomyopathy and isolated mitochondrial complex II deficiency

BACKGROUND Defects of the mitochondrial respiratory chain complex II (succinate dehydrogenase (SDH) complex) are extremely rare. Of the four nuclear encoded proteins composing complex II, only mutations in the 70 kDa flavoprotein (SDHA) and the recently identified complex II assembly factor (SDHAF1) have been found to be causative for mitochondrial respiratory chain diseases. Mutations in the other three subunits (SDHB, SDHC, SDHD) and the second assembly factor (SDHAF2) have so far only been associated with hereditary paragangliomas and phaeochromocytomas. Recessive germline mutations in SDHB have recently been associated with complex II deficiency and leukodystrophy in one patient. METHODS AND RESULTS We present the clinical and molecular investigations of the first patient with biochemical evidence of a severe isolated complex II deficiency due to compound heterozygous SDHD gene mutations. The patient presented with early progressive encephalomyopathy due to compound heterozygous p.E69 K and p.*164Lext*3 SDHD mutations. Native polyacrylamide gel electrophoresis and western blotting demonstrated an impaired complex II assembly. Complementation of a patient cell line additionally supported the pathogenicity of the novel identified mutations in SDHD. CONCLUSIONS This report describes the first case of isolated complex II deficiency due to recessive SDHD germline mutations. We therefore recommend screening for all SDH genes in isolated complex II deficiencies. It further emphasises the importance of appropriate genetic counselling to the family with regard to SDHD mutations and their role in tumorigenesis.

A multiplex PCR for the simultaneous detection and genotyping of the Echinococcus granulosus complex.

Echinococcus granulosus is characterized by high intra-specific variability (genotypes G1-G10) and according to the new molecular phylogeny of the genus Echinococcus, the E. granulosus complex has been divided into E. granulosus sensu stricto (G1-G3), E. equinus (G4), E. ortleppi (G5), and E. canadensis (G6-G10). The molecular characterization of E. granulosus isolates is fundamental to understand the spatio-temporal epidemiology of this complex in many endemic areas with the simultaneous occurrence of different Echinococcus species and genotypes. To simplify the genotyping of the E. granulosus complex we developed a single-tube multiplex PCR (mPCR) allowing three levels of discrimination: (i) Echinococcus genus, (ii) E. granulosus complex in common, and (iii) the specific genotype within the E. granulosus complex. The methodology was established with known DNA samples of the different strains/genotypes, confirmed on 42 already genotyped samples (Spain: 22 and Bulgaria: 20) and then successfully applied on 153 unknown samples (Tunisia: 114, Algeria: 26 and Argentina: 13). The sensitivity threshold of the mPCR was found to be 5 ng Echinoccoccus DNA in a mixture of up to 1 µg of foreign DNA and the specificity was 100% when template DNA from closely related members of the genus Taenia was used. Additionally to DNA samples, the mPCR can be carried out directly on boiled hydatid fluid or on alkaline-lysed frozen or fixed protoscoleces, thus avoiding classical DNA extractions. However, when using Echinococcus eggs obtained from fecal samples of infected dogs, the sensitivity of the mPCR was low (<40%). Thus, except for copro analysis, the mPCR described here has a high potential for a worldwide application in large-scale molecular epidemiological studies on the Echinococcus genus.

Syntheses, receptor bindings, in vitro and in vivo stabilities and biodistributions of DOTA-neurotensin(8-13) derivatives containing β-amino acid residues - a lesson about the importance of animal experiments

Neurotensin(8-13) (NTS(8-13)) analogs with C- and/or N-terminal β-amino acid residues and three DOTA derivatives thereof have been synthesized (i.e., 1-6). A virtual docking experiment showed almost perfect fit of one of the 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) derivatives, 6a, into a crystallographically identified receptor NTSR1 (Fig.1). The affinities for the receptors of the NTS analogs and derivatives are low, when determined with cell-membrane homogenates, while, with NTSR1-exhibiting cancer tissues, affinities in the single-digit nanomolar range can be observed (Table 2). Most of the β-amino acid-containing NTS(8-13) analogs (Table 1 and Fig.2), including the (68) Ga complexes of the DOTA-substituted ones (6; Figs.2 and 5), are stable for ca. 1 h in human serum and plasma, and in murine plasma. The biodistributions of two (68) Ga complexes (of 6a and 6b) in HT29 tumor-bearing nude mice, in the absence and in the presence of a blocking compound, after 10, 30, and 60 min (Figs. 3 and 4) lead to the conclusion that the amount of specifically bound radioligand is rather low. This was confirmed by PET-imaging experiments with the tumor-bearing mice (Fig.6). Comparison of the in vitro plasma stability (after 1 h) with the ex vivo blood content (after 10-15 min) of the two (68) Ga complexes shows that they are rapidly cleaved in the animals (Fig.5).

Widows may have fewer social and financial problems than in the past (Interview von Shereen Lehman mit Prof. Pasqualina Perrig-Chiello zu finanziellen und sozialen Problemen von Verwitweten)

(Reuters Health) – - A new Swiss study says that widows and widowers still mourn their spouses as much as ever, but compared to 35 years ago, everyday life is easier, especially for women. Widows, at least in Switzerland, have fewer financial troubles and more social connections than their counterparts in 1979, but widowers still complain of loneliness, researchers found.

A New Family of High-Affinity Transporters for Adenine, Cytosine, and Purine Derivatives in Arabidopsis

In many organisms, including plants, nucleic acid bases and derivatives such as caffeine are transported across the plasma membrane. Cytokinins, important hormones structurally related to adenine, are produced mainly in root apices, from where they are translocated to shoots to control a multitude of physiological processes. Complementation of a yeast mutant deficient in adenine uptake (fcy2) with an Arabidopsis cDNA expression library enabled the identification of a gene, AtPUP1 (for Arabidopsis thaliana purine permease1), belonging to a large gene family (AtPUP1 to AtPUP15) encoding a new class of small, integral membrane proteins. AtPUP1 transports adenine and cytosine with high affinity. Uptake is energy dependent, occurs against a concentration gradient, and is sensitive to protonophores, potentially indicating secondary active transport. Competition studies show that purine derivatives (e.g., hypoxanthine), phytohormones (e.g., zeatin and kinetin), and alkaloids (e.g., caffeine) are potent inhibitors of adenine and cytosine uptake. Inhibition by cytokinins is competitive (competitive inhibition constant Ki = 20 to 35 μM), indicating that cytokinins are transported by this system. AtPUP1 is expressed in all organs except roots, indicating that the gene encodes an uptake system for root-derived nucleic acid base derivatives in shoots or that it exports nucleic acid base analogs from shoots by way of the phloem. The other family members may have different affinities for nucleic acid bases, perhaps functioning as transporters for nucleosides, nucleotides, and their derivatives.