Hemin interactions and alterations of the subcellular localization of prion protein

Hemin (iron protoporphyrin IX) is a crucial component of many physiological processes acting either as a prosthetic group or as an intracellular messenger. Some unnatural, synthetic porphyrins have potent anti-scrapie activity and can interact with normal prion protein (PrPC). These observations raised the possibility that hemin, as a natural porphyrin, is a physiological ligand for PrPC. Accordingly, we evaluated PrPC interactions with hemin. When hemin (3-10 microM) was added to the medium of cultured cells, clusters of PrPC formed on the cell surface, and the detergent solubility of PrPC decreased. The addition of hemin also induced PrPC internalization and turnover. The ability of hemin to bind directly to PrPC was demonstrated by hemin-agarose affinity chromatography and UV-visible spectroscopy. Multiple hemin molecules bound primarily to the N-terminal third of PrPC, with reduced binding to PrPC lacking residues 34-94. These hemin-PrPC interactions suggest that PrPC may participate in hemin homeostasis, sensing, and/or uptake and that hemin might affect PrPC functions.     

Down-regulation of dopamine transporter by iron chelation in vitro is mediated by altered trafficking, not synthesis

Neurological development and functioning of dopamine (DA) neurotransmission is adversely affected by iron deficiency in early life. Iron-deficient rats demonstrate significant elevations in extracellular DA and a reduction in dopamine transporter (DAT) densities in the caudate putamen and nucleus accumbens. To explore possible mechanisms by which cellular iron concentrations control DAT functioning, endogenous DAT-expressing PC12 cells were used to determine the effect of iron chelation on DAT protein and mRNA expression patterns. In addition, we used human DAT (hDAT)-transfected Neuro2a (N2A) cells to examine DAT degradation and trafficking patterns. A 50 microM treatment for 24 h with the iron chelator, desferrioxamine (DFO), significantly decreased dopamine uptake in a dose-dependent manner, with no apparent change in K(m), in both PC12 and N2A cells. Reduced DA uptake was accompanied by concentration- and time-dependent reductions in total DAT protein levels in both cell lines. Exposure to increasing concentrations of DFO did not significantly alter DAT mRNA in either PC12 or N2A cells. However, DAT degradation rates increased three-fivefold in both cell types exposed to 50 microM DFO for 24 h. Biotinylation studies in N2A cells indicate a more dramatic loss of DAT in the membrane fraction, while OptiPrep fractionation experiments revealed an increase in lysosomal DAT with iron chelation. Inhibition of protein kinase C activation with staurosporin prevented the effect of iron chelation on DAT function, suggesting that in vitro iron chelation affects DAT primarily through the effects on trafficking rather than on synthesis.     

Carpal tunnel syndrome due to an atypical deep soft tissue leiomyoma: The risk of misdiagnosis and mismanagement

Background

Eccrine carcinoma is a quite rare malignant tumor that typically arises from a normal sweat gland and that features a rather high recurrence rate subsequent to simple excision. Given its rather poor response to adjuvant therapy, wide excision of the lesion with tumor-free margins may offer a reasonable chance for long-term control of this neoplasm.

Case presentation

Herein, we report on an unusual case of perianal eccrine carcinoma, initially presenting as a perianal abscess.

Conclusion

Even though eccrine carcinomas would appear to be rare, when dealing with recurrent skin tumors or recurrent perianal fistulas, the possibility of eccrine carcinoma should be considered by consulting clinicians.     

Banksia born to burn

? Historical evidence of recurrent fire in many of the world's biomes suggests that fire may have had profound evolutionary influences on their extant floras. However, the role of fire as a selective force in the origin and evolution of plant traits remains controversial. ? Using Bayesian Monte-Carlo-Markov-Chain procedures and calibration points from the fossil record, we generated a dated phylogeny for the iconic Australian genus Banksia, and reconstructed the evolutionary/chronological position of five putatively fire-related traits. ? The fire-dependent trait, on-plant seed storage (serotiny), and associated fire-enhancing trait, dead floret retention, co-originated with the first appearance of Banksia 60.8 million yr ago (Palaeocene). Whether nonsprouting or resprouting is ancestral was indeterminable, but the first banksias were nonclonal. Derived traits, such as dead leaf retention (fire-enhancing) and clonality (underground budbanks; fire-avoiding), first appeared 26-16 million yr ago (Miocene) with the onset of seasonal drought and thus more frequent fire, and culminated in dead florets/bracts completely covering the persistent fruits in some species. ? Thus, fire may have been a selective force in the very origin of Banksia 40 million yr before the onset of climate seasonality in the Miocene, and continued to have an impact on the direction of evolution, favouring traits consistent with adaptation to an increasingly (sometimes less) fire-prone environment.     

Application of magnetic particle tracking velocimetry to quadrupole magnetic sorting of porcine pancreatic islets

Magnetic isolation is a promising method for separating and concentrating pancreatic islets of Langerhans for transplantation in Type 1 diabetes patients. We are developing a continuous magnetic islet sorter to overcome the restrictions of current purification methods that result in limited yield and viability. In Quadrupole Magnetic Sorting (QMS) islets are magnetized by infusing superparamagnetic microbeads into islets' vasculature via arteries that serve the pancreas. The performance of the islet sorter depends on the resulting speed of the islets in an applied magnetic field, a property known as magnetophoretic mobility. Essential to the design and successful operation of the QMS is a method to measure the magnetophoretic mobilities of magnetically infused islets. We have adapted a Magnetic Particle Tracking Velocimeter (MPTV) to measure the magnetophoretic mobility of particles up to 1,000 μm in diameter. Velocity measurements are performed in a well-characterized uniform magnetic energy gradient using video imaging followed by analysis of the video images with a computer algorithm that produces a histogram of absolute mobilities. MPTV was validated using magnetic agarose beads serving as islet surrogates and subjecting them to QMS. Mobility distributions of labeled porcine islets indicated that magnetized islets have sufficient mobility to be captured by the proposed sorting method, with this result confirmed in test isolations of magnetized islets.     

Effect of transition metal ions on the fluorescence and Taq-catalyzed polymerase chain reaction of tricyclic cytidine analogs

We describe a simple method for real-time monitoring of matrix metalloproteinase-9 (MMP-9) collagenolytic activity for native triple helical collagen IV with a surface plasmon resonance (SPR) biosensor. The proteolytic activity of MMP-9 is measured as a decrease in the SPR signal resulting from the cleavage of collagen IV immobilized on the sensor surface. The kinetic parameters of full-length MMP-9 and its catalytic domain—catalytic constant (k_cat), association rate constant (k_a), and dissociation rate constant (k_d)—were estimated by the SPR method. The presence of sodium chloride and a nonionic detergent Brij-35 in a reaction solution led to the lower collagenolytic activity of MMP-9, whereas they suppressed the nonspecific interaction between MMP-9 and a cysteamine-modified chip. The comparison of kinetic parameters between MMP-9 and its catalytic domain revealed that the association constant of MMP-9 is much larger than that of the catalytic domain, suggesting that the interplay among hemopexin-like domain, fibronectin type II repeats motif, and linker region (O-glycosylated domain) plays an important role in recognizing collagen IV.