Discrimination between Francisella tularensis and Francisella-like endosymbionts when screening ticks by PCR

The presence of Francisella-like endosymbionts in tick species known to transmit tularemia poses a potential diagnostic problem for laboratories that screen tick samples by PCR for Francisella tularensis. Tick samples initially considered positive for F. tularensis based on standard 16S rRNA gene PCR were found to be positive only for Francisella-like endosymbionts using a multitarget F. tularensis TaqMan assay (ISFtu2, tul4, and iglC) and 16S rRNA gene sequencing. Specificity of PCR-based diagnostics for F. tularensis should be carefully evaluated with appropriate specimen types prior to diagnostic use.     

Microfluidic chips for in vivo imaging of cellular responses to neural injury in Drosophila larvae

With powerful genetics and a translucent cuticle, the Drosophila larva is an ideal model system for live imaging studies of neuronal cell biology and function. Here, we present an easy-to-use approach for high resolution live imaging in Drosophila using microfluidic chips. Two different designs allow for non-invasive and chemical-free immobilization of 3(rd) instar larvae over short (up to 1 hour) and long (up to 10 hours) time periods. We utilized these 'larva chips' to characterize several sub-cellular responses to axotomy which occur over a range of time scales in intact, unanaesthetized animals. These include waves of calcium which are induced within seconds of axotomy, and the intracellular transport of vesicles whose rate and flux within axons changes dramatically within 3 hours of axotomy. Axonal transport halts throughout the entire distal stump, but increases in the proximal stump. These responses precede the degeneration of the distal stump and regenerative sprouting of the proximal stump, which is initiated after a 7 hour period of dormancy and is associated with a dramatic increase in F-actin dynamics. In addition to allowing for the study of axonal regeneration in vivo, the larva chips can be utilized for a wide variety of in vivo imaging applications in Drosophila.     

Dental use wear in extinct lemurs: evidence of diet and niche differentiation

A new technique for molar use-wear analysis is applied to samples of all 16 species of extinct lemurs with known dentitions, as well as to a large comparative sample of extant primates. This technique, which relies on the light refractive properties of wear pits and scratches as seen under a standard stereoscopic microscope, has shown itself to be effective in distinguishing the diets of ungulates and extant primates. We draw dietary inferences for each of the 16 extinct lemur species in our database. There is a strong phylogenetic signal, with the Palaeopropithecidae showing use-wear signatures similar to those of the Indriidae; extinct lemurids (Pachylemur spp.) showing striking similarities to extant lemurids (except Hapalemur spp.); and Megaladapis showing similarities to Lepilemur spp. Only the Archaeolemuridae have dietary signatures unlike those of any extant lemurs, with the partial exception of Daubentonia. We conclude that the Archaeolemuridae were hard-object feeders; the Palaeopropithecidae were seed predators, consuming a mixed diet of foliage and fruit to varying degrees; Pachylemur was a fruit-dominated mixed feeder, but not a seed predator; and all Megaladapis were leaf browsers. There is no molar use wear evidence that any of the extinct lemurs relied on terrestrial foods (C4 grasses, tubers, rhizomes). This has possible implications for the role of the disappearance of wooded habitats in the extinction of lemurs.     

β-Adrenergic Receptors Mediate a Stress-Induced Decrease in IGF-II mRNA in the Rat Cerebellum

1. Exposure to a combined forced swimming–confinement stress resulted in a decrease in insulin-like growth factor II (IGF-II) mRNA levels in the whole brain (without the cerebellum) and in the isolated brain areas of the cerebral cortex, the hippocampus, and the cerebellum.2. In an effort to elucidate the neurotransmitter systems involved in this stress-induced decrease, animals were injected prior to exposure to the stress, with either propranolol, diazepam, or MK-801.3. Administration of diazepam or MK-801 did not affect the stress-induced decrease in IGF-II mRNA in any of the three brain areas examined.4. Administration of propranolol prior to the exposure to the stress inhibited the stress-induced decrease in IGF-II mRNA in the cerebellum. Propranolol had no such effect in the cerebral cortex or the hippocampus.5. Our results suggest that in the cerebellum, the stress-induced decrease in IGF-II mRNA is mediated by ₂-adrenergic receptors.     

Lumican regulates osteosarcoma cell adhesion by modulating TGFβ2 activity

Human osteosarcoma cell lines were recently shown to express and secrete the small leucine rich proteoglycan (SLRP) lumican, with the ability to regulate the growth and motility of these cells. In this study, lumican-deficient Saos 2 cells were demonstrated to have increased adhesive capability onto fibronectin (FN) (p≤0.01). Upon neutralization of endogenous transforming growth factor β2 (TGF-β2) activity, no difference in the ability of lumican siRNA-transfected and scramble siRNA-transfected Saos 2 cells to adhere onto FN was detected (p=NS). Exogenous TGF-β2 was shown to stimulate Saos 2 cell adhesion to FN (p≤0.01). These results therefore, suggest that the inverse correlation existing between lumican expression and Saos 2 cell adhesion is dependent on active TGF-β2 signaling. Furthermore, the significant increase in Smad 2 activation present in lumican-deficient cells (p≤0.01) was annulled in the presence of the anti-TGF-β2 peptide, demonstrating that lumican is an upstream regulator of the TGF-β2/Smad 2 signaling cascade. Crucial to FN-dependent adhesion, β1 integrin expression and pFAK activation were likewise identified as downstream TGF-β2 effectors regulated by lumican expression. In conclusion, this study demonstrates a novel out-in signaling circuit in human osteosarcoma cells: secreted to extracellular matrix lumican is an endogenous inhibitor of TGF-β2 activity, resulting in downstream effector modulation including pSmad 2, integrin β1 and pFAK to regulate osteosarcoma adhesion.     

Development of potent, orally active 1-substituted-3,4-dihydro-2-quinolone glycogen phosphorylase inhibitors

A series of substituted 3,4-dihydro-2-quinolone glycogen phosphorylase inhibitors, which have potential as antidiabetic agents, is described. Initial members of the series showed good enzyme inhibitory potency but poor physical properties. Optimisation of the 1-substituent led to 2,3-dihydroxypropyl compounds which showed good in vitro potency and improved physical properties, together with good DMPK profiles and acute in vivo efficacy in a rat model. X-ray crystallographic data are presented, showing an unexpected variety of binding orientations at the dimer interface site.