Cholesterol Levels Affect the Performance of AuNPs-Decorated Thermo-Sensitive Liposomes as Nanocarriers for Controlled Doxorubicin Delivery.

Stimulus-responsive liposomes (L) for triggering drug release to the target site are particularly useful in cancer therapy. This research was focused on the evaluation of the effects of cholesterol levels in the performance of gold nanoparticles (AuNPs)-functionalized L for controlled doxorubicin (D) delivery. Their interfacial and morphological properties, drug release behavior against temperature changes and cytotoxic activity against breast and ovarian cancer cells were studied. Langmuir isotherms were performed to identify the most stable combination of lipid components. Two mole fractions of cholesterol (3.35 mol% and 40 mol%, L1 and L2 series, respectively) were evaluated. Thin-film hydration and transmembrane pH-gradient methods were used for preparing the L and for D loading, respectively. The cationic surface of L allowed the anchoring of negatively charged AuNPs by electrostatic interactions, even inducing a shift in the zeta potential of the L2 series. L exhibited nanometric sizes and spherical shape. The higher the proportion of cholesterol, the higher the drug loading. D was released in a controlled manner by diffusion-controlled mechanisms, and the proportions of cholesterol and temperature of release media influenced its release profiles. D-encapsulated L preserved its antiproliferative activity against cancer cells. The developed liposomal formulations exhibit promising properties for cancer treatment and potential for hyperthermia therapy.

Host Suitability and Fitness-Related Parameters in Coptera haywardi (Hymenoptera: Diapriidae) Reared on Irradiated Ceratitis capitata (Diptera: Tephritidae) Pupae Stemming From the tsl Vienna-8 Genetic Sexing Strain.

Coptera haywardi (Ogloblin) is a pupal endoparasitoid of tephritid flies with great potential as a biological control agent worldwide as it does not attack other Diptera. To reach its full potential, its mass rearing needs to be enhanced lowering costs. Here, we focused on the use of irradiated pupae of Ceratitis capitata (Wiedemann) stemming from the temperature-sensitive lethal (tsl) Vienna-8 genetic sexing strain (= CcVienna-8), which is mass-produced in the San Juan Medfly and Parasitoid Mass Rearing Facility in Argentina. Exposure of 1- to 2-d-old CcVienna-8 pupae irradiated at 90 Gy to 6- to 8-d-old C. haywardi females at a 10:1 host/parasitoid ratio for 24 h turned out to be highly successful for the rearing of this parasitoid. High radiation doses (90-100 Gy) did not adversely influence fitness parameters of C. haywardi offspring F1, namely lifetime reproductive rates, adult life expectancy, and survival time. Demographic parameters in C. haywardi F1 from irradiated CcVienna-8 young pupae were improved compared to those values recorded from parasitoid originated from nonirradiated CcVienna-8 pupae. These findings will help to enhance parasitoid mass rearing for augmentative releases against medfly in Argentinean fruit-producing regions.

Her2p molecular modeling, mutant analysis and intramitochondrial localization.

Bacterial GatCAB amidotransferases are responsible for the transamidation of mischarged glutamyl-tRNA(Gln) into glutaminyl-tRNA(Gln). Mitochondria matrix also has a multienzymatic complex necessary for the transamidation of glutamyl-tRNA(Gln). Gtf1p, Her2p and Pet112p are the constituents of mitochondrial GatFAB amidotransferase complex. Her2p is subunit A of GatFAB complex, while Gtf1p is subunit F, a connector protein between Pet112p (subunit B) and Her2p. Here we evaluate through molecular modeling and amino acid correlation analysis the HER2 protein family. Localization studies indicated that Her2p is predominantly localized in the mitochondrial outer membrane, but it is also located in the mitochondrial matrix where together with Pet112p and Gtf1p constitutes the GatFAB complex. Finally, HER2 random mutagenesis unveiled important residues that provide thermo stability for the complex and are differently suppressed by overexpression of GTF1 or PET112. For instance, her2/ts11 mutant showed its fermentative growth impaired, and poorly rescued by GTF1 indicating that Her2p unknown function in the mitochondria outer membrane affects cell viability.

Clonagem, sequenciamento, expressão e caracterização do gene tsh de uma cepa de Escherichia coli patogênica de aves / Cloning, sequencing, expression, and characterization of the tsh gene from an avian pathogenic Escherichia coli strain

A hemaglutinina temperatura sensível (Tsh) pertence à família das serino-proteases autotransporte de Enterobacteriacea (SPATE), as quais são capazes de clivar diferentes substratos. Nós isolamos e caracterizamos o gene de Escherichia coli patogênica aviária (APEC) amostra APEC 13, sorotipo O2:H9, clonado em pET 101. A região de 4.2 kb do DNA clonado codificou uma proteína de aproximadamente 140 kDa (r-Tsh). O plasmídio recombinante pET 101-tsh conferiu um fenótipo de hemaglutinação positivo para a linhagem BL21 (tsh) para eritrócitos de galinha. A proteína r-Tsh foi purificada em coluna de níquel e utilizada na produção de anticorpos anti-Tsh. Um fragmento de 1.6 kb foi amplificado e subclonado em pCR4, e a seqüência parcial mostrou alta homologia com outras seqüências analisadas. O anti-Tsh reagiu com as proteínas r-Tsh e Tsh nativa da amostra APEC13, como demonstrado pela técnica de Western blot, mostrando que a r-Tsh tem epitopos conservados e que sua antigenicidade foi preservada. O anti-Tsh também inibiu a atividade hemaglutinante das amostras APEC 13 e BL12/pET 101-tsh

The temperature-sensitive hemagglutinin (Tsh) belongs to a family of high-molecular-weight serineprotease autotransporters of Enterobacteriaceae (SPATEs), which can cleave different substrates. Weisolated and characterised the tsh gene from an avian pathogenic Escherichia coli (APEC) strain, APEC13serotype O2:H9, which was cloned in pET101. The 4.2 kb region of cloned DNA coded one protein ofapproximately 140 kDa (r-Tsh). The recombinant plasmid pET101-tsh conferred to E. coli BL21 strain(tsh) the hemagglutination-positive phenotype against chicken erythrocytes. The r-Tsh was purified byNi-NTA column and used to produce antibody anti-Tsh. A 1.6 kb fragment of the tsh sequence was alsoamplified and cloned in pCR4, and a partial sequence showed high homology with other sequenceanalysed. The anti-Tsh reacted with the protein r-Tsh and native Tsh of APEC13, as demonstrated byWestern blot, showing that r-Tsh has conserved epitopes and that its antigenicity was preserved. Theanti-Tsh also inhibited the hemagglutinating activity of strains APEC13 and BL21/pET101-tsh

Cloning, sequencing, expression, and characterization of the tsh gene from an avian pathogenic Escherichia coli strain / Clonagem, sequenciamento, expressão e caracterização do gene tsh de uma cepa de Escherichia coli patogênica de aves

The temperature-sensitive hemagglutinin (Tsh) belongs to a family of high-molecular-weight serine protease autotransporters of Enterobacteriaceae (SPATEs), which can cleave different substrates. We isolated and characterised the tsh gene from an avian pathogenic Escherichia coli (APEC) strain, APEC13 serotype O2:H9, which was cloned in pET101. The 4.2 kb region of cloned DNA coded one protein of approximately 140 kDa (r-Tsh). The recombinant plasmid pET101-tsh conferred to E. coli BL21 strain (tsh) the hemagglutination-positive phenotype against chicken erythrocytes. The r-Tsh was purified by Ni-NTA column and used to produce antibody anti-Tsh. A 1.6 kb fragment of the tsh sequence was also amplified and cloned in pCR4, and a partial sequence showed high homology with other sequence analysed. The anti-Tsh reacted with the protein r-Tsh and native Tsh of APEC13, as demonstrated by Western blot, showing that r-Tsh has conserved epitopes and that its antigenicity was preserved. The anti-Tsh also inhibited the hemagglutinating activity of strains APEC13 and BL21/pET101-tsh.

A hemaglutinina temperatura sensível (Tsh) pertence à família das serino-proteases autotransporte de Enterobacteriacea (SPATE), as quais são capazes de clivar diferentes substratos. Nós isolamos e caracterizamos o gene de Escherichia coli patogênica aviária (APEC) amostra APEC 13, sorotipo O2:H9,clonado em pET101. A região de 4.2 kb do DNA clonado codifificou uma proteína de aproximadamente 140 kDa (r-Tsh). O plasmídio recombinante pET101-tsh conferiu um fenótipo de hemaglutinação positivo para a linhagem BL21 (tsh-) para eritrócitos de galinha. A proteína r-Tsh foi purificada em coluna de níquel e utilizada na produção de anticorpos anti-Tsh. Um fragmento de 1.6 kb foi amplificado e subclonado em pCR4, e a seqüência parcial mostrou alta homologia com outras seqüências analisadas. O anti-Tsh reagiu com as proteínas r-Tsh e Tsh nativa da amostra APEC13, como demonstrado pela técnica de Western blot, mostrando que a r-Tsh tem epitopos conservados e que sua antigenicidade foi preservada. O anti-Tsh também inibiu a atividade hemaglutinante das amostras APEC13 e BL21/pET 101-tsh.