Radioiodinated azide and isothiocyanate derivatives of cocaine for irreversible labeling of dopamine transporters: synthesis and covalent binding studies

Two novel N-substituted-3beta-phenyltropane alkaloids have been labeled with iodine-125 for use as irreversible probes of dopamine transporter (DAT) binding sites. One contains an iodoaryl azide moiety for photolabeling, while the other bears an iodoaryl isothiocyanate for direct conjugation. Both radioligands were prepared in a one-flask procedure by electrophilic radioiodination of the corresponding aniline under no-carrier-added conditions, followed either by diazotization and treatment with sodium azide, or by addition of thiophosgene under basic conditions. Specifically, (-)-N-[4-(3-[(125)I]iodo-4-azidophenyl)butyl]-2beta-carbomethoxy-3beta-(4-chlorophenyl)tropane ([(125)I]MFZ-2-24) and (-)-N-[4-(3-[(125)I]iodo-4-isothiocyanophenyl)butyl]-2beta-carbomethoxy-3beta-(4-chlorophenyl)tropane ([(125)I]MFZ 3-37) were synthesized. Isolation by reversed-phase HPLC and solid-phase extraction gave good average yields of [(125)I]MFZ-2-24 (67%, n = 5) and [(125)I]MFZ-3-37 (45%, n = 3) with high radiochemical purities (96-99%) and specific radioactivities (>2000 mCi/micromol). The utility of the radioligands was demonstrated by their covalent linkage to rat striatal membranes, and immunoprecipitation of a single radiolabeled band at 80 kDa corresponding to the full-length DAT.     

The mitogen-activated protein kinase kinase kinase dual leucine zipper-bearing kinase (DLK) acts as a key regulator of keratinocyte terminal differentiation

In the skin, epithelial cells undergo a terminal differentiation program leading to the formation of the stratum corneum. Although it is expected that the last phases of this process must be tightly regulated since it results in cell death, the signaling pathways involved in this induction remain ill defined. We now report that a single kinase, the mitogen-activated protein kinase kinase kinase dual leucine zipper-bearing kinase (DLK), acts in the epidermis to promote the terminal differentiation of human keratinocytes. In support of this notion, we showed that DLK expression was restricted to the granular layer in situ. In addition, cultured keratinocytes infected with a recombinant adenovirus expressing DLK exhibited morphological and biochemical changes, including a suprabasal localization, altered cell shape, compacted cytoplasm, DNA fragmentation, and the up-regulation of filaggrin, that are reminiscent of a terminally differentiated phenotype. Moreover the expression of wild-type DLK in keratinocytes stimulated transglutaminase activity and the consequent formation of the cornified cell envelope, while a kinase-inactive variant of DLK did not. Together these results identify DLK as a signaling molecule implicated in the regulation of keratinocyte terminal differentiation and cornification.     

Practical Methodology of Cognitive Tasks Within a Navigational Assessment

This paper describes an approach for measuring navigation accuracy relative to cognitive skills. The methodology behind the assessment will thus be clearly outlined in a step-by-step manner. Navigational skills are important when trying to find symbols within a speech-generating device (SGD) that has a dynamic screen and taxonomical organization. The following skills have been found to impact children’s ability to find symbols when navigating within the levels of an SGD: sustained attention, categorization, cognitive flexibility, and fluid reasoning^1,2. According to past studies, working memory was not correlated with navigation^1,2.The materials needed for this method include a computerized tablet, an augmentative and alternative communication application, a booklet of symbols, and the Leiter International Performance Scale-Revised (Leiter-R)³. This method has been used in two previous studies. Robillard, Mayer-Crittenden, Roy-Charland, Minor-Corriveau and Bélanger¹ assessed typically developing children, while Rondeau, Robillard and Roy-Charland² assessed children and adolescents with a diagnosis of Autism Spectrum Disorder. The direct observation of this method will facilitate the replication of this study for researchers. It will also help clinicians that work with children who have complex communication needs to determine the children’s ability to navigate an SGD with taxonomical categorization.     

Diverse Genotypes of Yersinia pestis Caused Plague in Madagascar in 2007

Background Yersinia pestis is the causative agent of human plague and is endemic in various African, Asian and American countries. In Madagascar, the disease represents a significant public health problem with hundreds of human cases a year. Unfortunately, poor infrastructure makes outbreak investigations challenging.Conclusions/SignificancePlague in Madagascar is caused by numerous distinct types of Y. pestis. Genotyping method choice should be based upon the discriminatory power needed, expense, and available data for any desired comparisons. We conclude that genotyping should be a standard tool used in epidemiological investigations of plague outbreaks.     

Age‐associated expression of p21and p53 during human wound healing

In mice, cellular senescence and senescence‐associated secretory phenotype (SASP) positively contribute to cutaneous wound healing. In this proof‐of‐concept study, we investigated the expressions of p16, p21, and other senescence‐associated biomarkers during human wound healing in 24 healthy subjects using a double‐biopsy experimental design. The first punch biopsy created the wound and established the baseline. The second biopsy, concentric to the first and taken several days after wounding, was used to probe for expression of biomarkers by immunohistochemistry and RNA FISH. To assess the effects of age, we recruited 12 sex‐matched younger (30.2 ± 1.3 years) and 12 sex‐matched older (75.6 ± 1.8 years) subjects. We found that p21 and p53, but not p16, were induced during healing in younger, but not older subjects. A role for Notch signaling in p21 expression was inferred from the inducible activation of HES1. Further, other SASP biomarkers such as dipeptidyl peptidase‐4 (DPP4) were significantly induced upon wounding in both younger and older groups, whereas matrix metallopeptidase 9 (MMP9) was induced only in the younger group. Senescence‐associated β‐galactosidase (SA‐β‐gal) was not detectable before or after wounding. This pilot study suggests the possibility that human cutaneous wound healing is characterized by differential expression of p21 and p53 between younger and older subjects.     

Amino acid signaling to mTOR mediated by inositol polyphosphate multikinase

mTOR complex 1 (mTORC1; mammalian target of rapamycin [mTOR] in complex with raptor) is a key regulator of protein synthesis and cell growth in response to nutrient amino acids. Here we report that inositol polyphosphate multikinase (IPMK), which possesses both inositol phosphate kinase and lipid kinase activities, regulates amino acid signaling to mTORC1. This regulation is independent of IPMK's catalytic function, instead reflecting its binding with mTOR and raptor, which maintains the mTOR-raptor association. Thus, IPMK appears to be a physiologic mTOR cofactor, serving as a determinant of mTORC1 stability and amino acid-induced mTOR signaling. Substances that block IPMK-mTORC1 binding may afford therapeutic benefit in nutrient amino acid-regulated conditions such as obesity and diabetes.     

Complete genome sequence of Treponema succinifaciens type strain (6091)

Treponema succinifaciens Cwyk and Canale-Parola 1981 is of interest because this strictly anaerobic, apathogenic member of the genus Treponema oxidizes carbohydrates and couples the Embden-Meyerhof pathway via activity of a pyruvate-formate lyase to the production of acetyl-coenzyme A and formate. This feature separates this species from most other anaerobic spirochetes. The genome of T. succinifaciens 6091(T) is only the second completed and published type strain genome from the genus Treponema in the family Spirochaetaceae. The 2,897,425 bp long genome with one plasmid harbors 2,723 protein-coding and 63 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.